Revision as of 18:26, 16 September 2015

Dundee iGEM 2015

LABJOURNAL

BioSpray

Our forensic toolkit aims to use synthetic biology approaches to improve on the current methods used by crime scene investigators.

Chromium Detector

Lorem ipsum dolor sit amet, consectetur adipiscing elit. Curabitur a tincidunt elit. Aliquam porta nibh at enim luctus, auctor consequat dolor vehicula.

Fingerprint Aging

Lorem ipsum dolor sit amet, consectetur adipiscing elit. Curabitur a tincidunt elit. Aliquam porta nibh at enim luctus, auctor consequat dolor vehicula.

Week Beginning 22/06/2015

Summary

This week was all about getting started with the newly arrived biobrick BBa_K1058008.

Day 1 - 22/06: Plated BBa_K1058008 from agar stab.

Aim of experiment: To prepare BBa_K1058008 for sequence confirmation, storage, and further processing. The culture was plated from the agar stab and also a liquid culture set up.

Protocols Used:

Click here to see our plating protocol.

Click here to see our overnight culture protocol.

Results: N/A

Next Steps: Purification of the Plasmid, containing BBa_K1058008 and confirmation of size and sequence.

Day 2 - 23/06: Purified BBa_K1058008 from liquid culture.

Aim of experiment: To purify BBa_K1058008 for sequence confirmation.

Protocols Used: Miniprep plasmid purification, DNA sequencing

Results Sequencing: Figure 1

Next Steps: PCR of promoter region (pChr) and repressor region (ChrB) of BBa_K1058008 for storage in individual biobricks.

Week Beginning 29/06

Summary

During this week, BBa_K1058008 was disassembled into its constituent parts, pChr, and ChrB.

Day 1 - 29/06: PCR-amplification of pChr and ChrB from BBa_K1058008.

Aim of experiment: To break down BBa_K1058008 into the promoter region pChr, and the repressor gene, ChrB, while adding standard prefix and suffix to each. Furthermore optimisation of first 15 codons of ChrB in order to increase the overall rate of transcription. Restriction digest of the respective ends and subsequent ligation of pChr and ChrB into pSB1C3 for submission to registry.

Protocols Used: PCR, Gel extraction, Restriction Digest

Results: Figure 2

Next Steps: Repeat of PCR of ChrB. Purification of the Plasmid, containing BBa_K1058008 and confirmation of size and sequence.

Week Beginning 06/07

Summary

During this week ligations of pChr and ChrB into pSB1C3 were finalised and the constituent parts of our newly designed chromate sensing system were prepared for ligation and transformation.

Day 1 - 06/07: Repeated amplification of ChrB. Ligation and transformation of pChr and ChrB.

Aim of experiment: To repeat PCR of ChrB for subsequent ligation into pSB1C3. Ligation of pChr and ChrB into pSB1C3. Transformation of each into a non-pathogenic lab strain of Escherichia coli. Overnight culture of E. coli DH5-alpha, containing GFPmut2, as a fluorescent reporter for our chromate sensing system

Protocols Used: PCR-amplification, Gel extraction, Restriction digest, Ligation, Overnight cultures

Results: Figure 3

2 bands were extracted from the gel for further characterisation, the strongest band, hence called ChrBs, and the band directly above, hence called ChrBw. pChr, ChrBs, and ChrBw were ligated into pSB1C3. pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw were transformed into E. coli MC1061.

Next Steps: Sequence and size confirmation of pChr-pSB1C3, ChrBs-pSB1C3, and ChrBw-pSB1C3. Ligation of pChr to GFP coding sequence for construction of a reporter plasmid.

Day 2 - 07/07: Purification of GFP and overnight cultures of pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw

Aim of experiment: Purification of GFPmut2 from overnight cultures. Overnight cultures of recombinant E. coli MC1061, containing pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw respectively for further size and sequence confirmation. Overnight culture of pUniprom for subsequent ligation of ChrB

Protocols Used: Miniprep, PCR-amplification, Gel extraction, Overnight cultures

Results: Figure 5

Next Steps: Repeat of amplification of ChrB. Sequence and size confirmation of pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw.

Day 3 - 08/07: Purification of plasmids pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw. Size and sequence confirmation.

Aim of experiment: To purify overnight cultures for size and sequence confirmation. Repeat of PCR amplification of pChr and ChrB. Restriction digestes of all parts in order to produce compatible sticky ends.

Protocols Used: Miniprep, PCR amplification, Restriction digest, Sequencing

Restriction digest of gel purified PCR-products of pChr, ChrB, and GFPmut2, using EcoRI/SpeI, BamHI/HindIII, and XbaI/PstI respecitvely.

Results:

Figure 6 - Size confirmation of pChr and ChrB.

Figure 7 Sequence confirmation of pChr

Figure 8 Sequence confirmation of ChrB.

Next Steps: Purification and digest of pUniprom for insertion of ChrB.

Day 4 - 09/07: Digested and cleaned up pUniprom for insertion of ChrB. Amplification of optimised ChrB with primers specific for the insertion into pUniprom.

Aim of experiment: To prepare pUniprom for insertion of ChrB by digesting it with BamHI and HindIII. To amplify ChrB version with first 15 codons optimised for insertion into pUniprom.

Protocols Used:

Restriction digest and Gel extraction of pUniprom

PCR and Gel extraction of optimised ChrB

Results:

Figure 9 pUniprom

Figure 10 ChrB

Next Steps: Ligation of ChrB and codon optimised ChrB into pUniprom.

Day 5 - 10/07: Ligations of ChrB and optimised ChrB into pUniprom, and of pChr and GFPmut2 into pSB1C3.

Aim of experiment: Ligation of ChrB and optimised ChrB into pUniprom via BamHI and HindIII restriction sites. Ligation of pChr and GFPmut2 to each other via SpeI/XbaI-link and into pSB1C3 via EcoRI and PstI restriction sites.

Protocols Used: Ligations

Results: N/A

Next Steps: Transformation of abovementioned ligations.

Week Beginning 13/07

Summary

During this week successful transformants were identified and propagated.

Day 1 - 13/07: Transformation of pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP.

Aim of experiment: Transformation of pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP into E. coli JM110 chassis. Setting up a backup restriction digest of pSB1C3 with EcoRI and SpeI in case the double ligation pSB1C3-pChr-GFP is unsuccessful.

Protocols Used: Transformations, Restriction digest

Results: Figure 11

Next Steps: Purification of recombinant vectors and confirmation of size and sequence. Store digested pSB1C3 at -20^oC.

Day 2 - 14/07: Set up cultures for subsequent purification of recombinant vectors for size and sequence confirmation.

Aim of experiment: Purification of recombinant vectors, pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP, for size and sequence confirmation. Patch plating of the same colonies for storage and retrieval once size and sequence have been confirmed.

Protocols Used: Overnight cultures, Patch plating

Results: N/A

Next Steps: Purification of the recombinant plasmids for confirmation of size and sequence.

Day 3 - 15/07: Purified pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP from overnight cultures

Aim of experiment: Purification of pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP from overnight for confirmation of size and sequence. Restriction digest of pUnipromChrB and optimised pUniprom-ChrB optimised with BamHI and HindIII and of pSB1C3-pChr-GFP with EcoRI/PstI for size confirmation.

Protocols Used: Miniprep, Restriction digest, Colony PCR, Sequencing

Results:

Figure 12 Size confirmation of ChrB and ChrBopt.

Figure 13 Gel after colony PCR. Colonies 2, 4, 9, and 12 were selected for sequencing.

Figure 14 Sequencing result of pSB1C3-pChr-GFPmut2_1. The Result for pSB1C3-pChr-GFPmut2_2 was not as expected, albeit in the correct orientation.

Next Steps: Confirmation of sequence of ChrB-opt.

Day 4 - 16/07: Set up overnight cultures of colonies chosen after colony-PCR for subsequent purification and sequence confirmation.

Aim of experiment: Set up overnight cultures of ChrBopt colonies 2, 4, 9, and 12 for sequencing.

Protocols Used: Overnight cultures

Results:N/A

Next Steps: Purification of recombinant plasmids.

Day 5 - 17/07: Purified and sequenced chosen ChrBopt-colonies.

Aim of experiment: To purify recombinant plasmids for sequence confirmation.

Protocols Used: Miniprep, Sequencing

Results: ChrBopt showed HindIII restriction sites and was hence cleaved into partial inserts.

Next Steps: Use alternative cloning protocol for ChrB and ChrBopt.

Week Beginning 20/07

Summary

During this week an alternative cloning strategy for ChrB and ChrBopt was employed. Furthermore, the pSB1C3-pChr-GFP construct was cloned in a stepwise fashion.

Day 1 - 20/07: Went back to the original plate of pSB1C3-pChr-GFP for visualising colonies that do or do not express GFP.

Aim of experiment:To compare colonies on the patch plate of pSB1C3-pChr-GFP by differentially visualising colonies that do or do not express GFP under UV light. The background of this experiment was that if there is one colony that has both inserts in the correct orientation with respect to each other, then there is perhaps one that also has them with the correct orientation with respect to the plasmid. Colonies identified as expressing GFP were patch plated and grown in liquid cultures over night in order to confirm the sequence of the insert.

Protocols Used: Patch-plating, overnight cultures

Results:Figure 15 pSB1C3-pChr-GFP in E. coli JM110.

Next Steps: Sequence confirmation of newly selected colonies.

Day 2 - 21/07: Purified plasmids from new colonies and confirmed sequence. Restriction digest of pUniprom with BamHI and PstI.

Aim of experiment:Purification of newly selected colonies, 1, 5, 6, and 7, containing pSB1C3-pChr-GFP for subsequent sequence confirmation. Restriction digest of pUniprom with BamHI and PstI to employ an alternative cloning strategy of ChrB and ChrBopt.

Protocols Used:

Miniprep and Sequencing of pChrGFP insert.

Restriction digest of pUniprom.

Results:

Figure 16 Alignment of sequencing results.

Figure 17 Gel after restriction digest of pUniprom.

Next Steps:Stepwise ligation of pChr and GFP into pSB1C3 in order to clone the inserts in the correct orientation with respect to the plasmid. Insertion of ChrB and ChrBopt into pUniprom via BamHI and PstI links.

Day 3 - 22/07: Ligated pChr into pSB1C3 via EcoRI and SpeI sites and cloned the recombinant plasmid into E. coli JM110.

Aim of experiment:Ligation of pChr into pSB1C3 via EcoRI SpeI link. This is the first step of a 2-step cloning strategy with the aim of cloning pChr and GFP into pSB1C3 in the correct orientation with respect to the plasmid.

Protocols Used: Ligation, Transformation

Results: Figure 18 Plates after transformation of pChr into pSB1C3.

Next Steps: Sequence confirmation of pChr in pSB1C3.

Day 4 - 23/07: Set up overnight cultures of pSB1C3-pChr colonies.

Aim of experiment: Set up overnight cultures of pSB1C3-pChr colonies to prepare pSB1C3-pChr for sequence confirmation

Protocols Used: Overnight culture, Patch-plating

Results: N/A

Next Steps: Purification of the recombinant vector for size and sequence confirmation.

Day 5 - 24/07: Purified pSB1C3-pChr for sequence confirmation.

Aim of experiment: Purification of pSB1C3-pChr for sequence confirmation before ligating GFP into it.

Protocols Used:

Plasmid purification, Sequencing

Results: Figure 19 Alignment of sequenced pSB1C3-pChr vectors with pChr sequence.

Next Steps: Digest of pSB1C3-pChr with SpeI/PstI and ligation of GFPmut2.

Day 6 - 25/07: PCR-amplified ChrB and ChrBopt.

Aim of experiment: Amplification of ChrB and ChrBopt for insertion into pUniprom. Restriction digest of the PCR product with BamHI and PstI for producing compatible sticky ends

Protocols Used: PCR-amplification, Restriction digest

Results: N/A

Next Steps:Ligation and Transformation of ChrB and ChrBopt.

Day 7 - 26/07: Ligated ChrB and ChrBopt into pUniprom.

Aim of experiment: Insertion of ChrB and optimised ChrB into pUniprom vector.

Protocols Used: Ligation

Results: N/A

Next Steps: Transformation of ligated vectors.

Week Beginning 27/07

Summary

Final touches were made on the chromate system. All parts were ligated into their final vectors during this week. Preparations were made for characterisation of each of the parts.

Day 1 - 27/07: Transformed ChrB and ChrBopt into E. coli JM110

Aim of experiment:Transformation of the repressors, ChrB and ChrBopt, into JM110

Protocols Used: Transformation

Results: Figure 20 Plates after transformation.

Next Steps: Purification of pUniprom-ChrB and pUniprom-ChrBopt for sequence confirmation.

Day 2 - 28/07: Digested pSB1C3-pChr for insertion of GFP

Aim of experiment:Digesting sequence confirmed pSB1C3-pChr with SpeI and PstI for subsequent ligation with GFP. Overnight cultures of ChrB and ChrBopt for subsequent plasmid purification, size confirmation, and sequence confirmation.

Protocols Used:

Restriction digest, Ligation of GFP into pSB1C3-pChr.

Overnight culture of of ChrB and ChrBopt.

Next Steps: Transformation of recombinant vector for confirmation of size and sequence.

Results: N/A

Next Steps:Transformation of pSB1C3-pChr-GFP for size and sequence confirmation. Purification of ChrB and ChrBopt for sequence confirmation.

Day 3 - 29/07: Purified ChrB and ChrBopt for size and sequence confirmation.

Aim of experiment: Purification of ChrB and ChrBopt for confirmation of size and sequence.

Protocols Used: Plasmid purification, Restriction digest, Sequencing

Results:

Figure 21 Confirmation of insert size.

Figure 22 Confirmation of sequence.

Next Steps: Confirmation of expression of ChrB and ChrBopt by western blotting.

Day 4 - 30/07: Repeated transformation of pSB1C3-pChr-GFP into E.coli JM110

Aim of experiment:Digest of pSB1C3-pChr with SpeI and PstI for insertion of GFP. Repeat of unsuccessful transformation from 28/07.

Protocols Used: Restriction digest, Ligation, Transformation

Results: Figure 23 Plates after transformation

Next Steps: Overnight cultures for purification of the pSB1C3-pChr-GFP for confirmation of insert size and sequence.

Day 5 - 31/07: Set up overnight cultures for confirmation of pSB1C3-pChr-GFP.

Aim of experiment: Overnight cultures for subsequent plasmid purification of pSB1C3-pChr-GFP.

Protocols Used: Overnight culture

Results: N/A

Next Steps: Purification of the Plasmids for confirmation of insert size and sequence.

Day 6 - 01/08: Purified pSB1C3-pChr-GFP for size confirmation and sequencing.

Aim of experiment: Purification of pSB1C3-pChr-GFP for confirmation of insert size and sequence.

Protocols Used: Plasmid purification, Restriction digest, Sequencing

Results:

Figure 24 Gel for size confirmation.

Figure 25 Sequence confirmation.

Next Steps:Overnight cultures in preparation for western blotting for characterisation of expression of both, GFP, and ChrB and ChrBopt.

Day 7 - 02/08: Set up cultures in preparation for western blotting.

Aim of experiment:Overnight cultures of sequence confirmed ChrB, ChrBopt, pChrGFP1 (from 15/07), pChrGFP3 and pChrGFP4 (31/07) for subsequent western blotting.

Protocols Used: Overnight culture

Results: N/A

Next Steps: Western blotting for characterisation of expression of both, GFP, and ChrB and ChrBopt.

Week Beginning 03/08

Summary

This week was entirely used for characterising the expression of GFP from the promoter pChr, and the expression of ChrB from the tat-promoter in pUniprom.

Day 1 - Day 7 - 03/08 - 09/08: Determined expression of ChrB, ChrBopt, and GFP by western blotting.

Aim of experiment:To determine whether or not the required proteins, GFP, and ChrB, are expressed by blotting against GFP and against a 6His-tag respectively.

Protocols Used: SDS-PAGE and western blot

Results: This procedure was repeated several times during this week in order to find appropriate concentrations and volumes to load. Figure 26 was blotted after SDS-PAGE with the following concentrations:

GFP: Dilute pellet of 1ml overnight culture with 1ml TBS. Take 100µl of this solution and mix with 100µl Laemmli Buffer. Load 3µl on the SDS-gel.
HIS: Dilute pellet of 1ml overnight culture with 200µl Laemmli buffer. Load 10µl on the SDS-gel.

Figure 26

Next Steps:Transformation of both plasmids into MG1655 with the goal to characterise the interaction between ChrB and pChr. Furthermore the original biobrick, BBa_K1058008 will be tested under the same conditions for comparison.

Week Beginning 10/08

Summary

This week was spent with transforming all combinations of pSB1C3-pChr-GFP and pUniprom-ChrB into E. coli MG1655. Tests for their interaction were also prepared.

Day 1 - 10/08: Transformed both pSB1C3-pChr-GFP-plasmids, pUniprom-ChrB, and pUniprom-ChrBopt, as well as BBa_K1058008 into E. coli MG1655.

Aim of experiment: To transform the following combinations of pSB1C3-pChr-GFP and pUniprom-ChrB into MG1655:

ChrB2 (29/07) - pChrGFP (31/07)
ChrB2 (29/07) - pChrGFP (15/07)
ChrBopt3 (29/07) - pChrGFP (31/07)
ChrBopt3 (29/07) - pChrGFP (15/07)

in a stepwise fashion. Furthermore transform BBa_K1058008 into MG1655. Transformations were done in a stepwise fashion with only one of the plasmids at a time.

Protocols Used: Plasmid purification, Transformation

Results: Figure 27

Next Steps:Making newly transformed cells competent again in order to transform the second plasmid as well.

Day 2 - 11/08: Set up overnight cultures of all transformed cells.

Aim of experiment: To prepare overnight cultures of yesterdays transformaions in order to make them competent to transform the second plasmid as well.

Protocols Used: Overnight culture

Results: N/A

Next Steps:Make cells of overnight culture competent to transform in second plasmid.

Day 3 - 12/08: Made competent MG1655 with pUniprom-ChrB2, pUniprom-ChrBopt3, pSB1C3-pChr-GFP (15/07), pSB1C3-pChr-GFP (31/07).

Aim of experiment:To make transformed cells competent in order to transform the second plasmid. The same plasmids that were used for the initial transformations were also used for this set of transformations.

Protocols Used: Competent cells, Transformations

Results: N/A

Next Steps:Blot against GFP in order to confirm (or refute) that ChrB interacts with pChr and hence switches off expression of GFP.

Day 4 - 13/08: Transformed pUniprom-ChrB2 and pUniprom-ChrBopt3 into MG1655+pSB1C3-pChr-GFP(15/07) and MG1655+pChrGFP(31/07)

Aim of experiment: To build a strain of E. coli MG1655, containing both plasmids required for a chromate sensor.

Protocols Used: Transformation

Results: No successful transformants were found.

Next Steps: Repeat this experiment but transform pSB1C3-pChr-GFP (15/07) and pSB1C3-pChr-GFP (31/07) into competent MG1655+pUniprom-ChrB2 and MG1655+pUniprom-ChrBopt3 respectively.

Week Beginning 17/08

Summary

During this week E. coli MG1655 containing both plasmids for a. Western Blots were used to determine expression of ChrB, ChrBopt, and GFP.

Day 1 - 17/08: Transformations akin to 13/08 were done, but pSB1C3-pChr-GFP(15/07) and pSB1C3-pChr-GFP(31/07) were transformed into MG1655+pUniprom-ChrB2 and MG1655+pUniprom-ChrBopt3.

Aim of experiment: To build a strain of MG1655, containing both plasmids required for a chromate sensor.

Protocols Used: Transformation

Results: Successful transformants were found on all plates.

Next Steps: To determine expression of ChrB and ChrBopt (via 6His-tag) and expression of GFP.

Day 2 - 18/08: Preparation of samples for western blot.

Aim of experiment: To determine whether ChrB, ChrBopt, and GFP are expressed and have first estimates of differences in expression levels.

Protocols Used: Overnight culture

Results: N/A

Next Steps: Lyse cells and prepare samples for SDS-PAGE + Western-Blot.

Day 3-6 - 19/08 - 22/08: Repeated western blots with different concentrations to determine whether the expression of ChrB has an effect on the expression of GFP.

Aim of experiment: To determine whether expression of ChrB or ChrBopt has an effect on the expression of GFP.

Protocols Used: SDS-PAGE + Western Blot

Results: Figure 28

Next Steps: Check if different chromate salts have an effect on the respective expression levels.

Week Beginning 24/08

Summary

Plate reader experiments were carried out in order to determine the effect of K₂CrO₄, K₂Cr₂O₇, CrCl₃, and K₂SO₄ on the expression of GFP.

Day 1 - Day 7 - 24/08-30/08: Design of Plate reader experiments.

Aim of experiment: To design a series of plate reader experiments to measure OD₆₀₀ and relative fluorescence of the chromate sensor as a response to different concentrations of K₂CrO₄, K₂Cr₂O₇, CrCl₃, and K₂SO₄.

Protocols Used: Plate Reader Experiments

Results: Figure 29 MG1655 + BBa_K1058008

Results: Figure 30 MG1655 + pSB1C3-pChr-GFP + pUniprom-ChrB

Results: Figure 31 MG1655 + pSB1C3-pChr-GFP + pUniprom-ChrBopt

Results: Figure 32 MG1655 + pSB1C3-pChr-GFP

Next Steps: Try the system with different expression levels of ChrB and ChrBopt. Induce overexpression by exploiting the T7-promoter in pUniprom after transforming the different combinations between pSB1C3-pChr-GFP and pUniprom-ChrB/pUniprom-ChrBopt into BL21(DE3) and adding IPTG.

Week Beginning 31/08

Summary

UK Synbio Conference and UK iGEM meetup at Westminster University.

Week Beginning 07/09

Summary

Parts from CeBiTec Bielefeld arrived and attempts at cloning were made. The chromate sensing system was transformed into BL21(DE3) and some of the plate reader experiments repeated.

Day 1 - 07/09: Transformation of pSB1K3 (Bielefeld) into MG1655.

Aim of experiment: The high-copy plasmid arrived at a concentration of 7 ^ng/_µl. Hence it was transformed in order to produce more of it before purifying and digesting it.

Protocols Used: Transformation,

Overnight culture for new E. coli MG1655

Results: No transformants were found.

Next Steps: Retry transformation with new competent E. coli MG1655.

Day 2 - 08/09: Transformation of pSB1K3 into MG1655.

Aim of experiment: Test transformation of BBa_K1058008 into new MG1655 competent cells and re-transformation of pSB1K3 for subsequent miniprep.

Protocols Used: Transformation

Results: Successful test transformation, but no colonies on pSB1K3 plate.

Next Steps: Use an alternative compatible plasmid for ligation of codon optimised ChrB (Bielefeld).

Day 3 - 09/09: Transformation of pSB1K3 into MG1655.

Aim of experiment: Restriction digest of pT7KS with EcoRI and PstI as alternative to pSB1K3. Ligation of digested ChrB (Bielefeld) and transformation of pT7KS-ChrB. Also transformations of Chromate-sensor plasmids into BL21 (DE3) in the following combinations:

pSB1C3-pChr-GFP + pUniprom-ChrB
pSB1C3-pChr-GFP + pUniprom-ChrBopt
pUniprom-ChrB
pUniprom-ChrBopt

Protocols Used: Restriction Digest, Gel Extraction, Ligation

Results:Transformations of the plasmid with pT7KS-ChrB (Bielefeld) yielded 12 colonies, but also colonies on the vector control plate. Transformations of pSB1C3-pChr-GFP + pUniprom-ChrB, pSB1C3-pChr-GFP + pUniprom-ChrBopt, pUniprom-ChrB, and pUniprom-ChrBopt into BL21(DE3) were successful.

Results:

Figure 33 pT7KS-ChrB (Bielefeld) transformations

Figure 34 Transformations into BL21(DE3)

Next Steps: Confirmation of colonies with pT7KS-ChrB by colony PCR. Preparation of recombinant BL21(DE3) for plate reader experiments.

Day 4 - 10/09: Colony PCR of pT7KS-ChrB (Bielefeld).

Aim of experiment: Check if any of the colonies after ligation of ChrB (Bielefeld) into pT7KS were successful despite colonies on the control plate.

Protocols Used: Colony PCR

Results: Figure 34

Next Steps: Prepare transformed BL21(DE3) for plate reader experiments. Attempt new transformation of pT7KS-ChrB (Bielefeld).

Day 5 - 11/09: Preparation for Plate reader experiment with BL21(DE3)

Aim of experiment: Set up overnight cultures for subsequent plate reader experiments.

Protocols Used: Overnight cultures

Results: N/A

Next Steps: Set up plate reader, collect and analyse data. Assess the effect of induction of higher repressor-expression on the level of GFP.

Day 6 - 12/09: Set up plate reader.

Aim of experiment: To design a series of plate reader experiments to measure OD₆₀₀ and relative fluorescence of the chromate sensor. At first no chromate was added to this plate in order to assess the effect of IPTG induction.

Protocols Used: Plate reader experiments

Results: Figure 35

Next Steps:Repeat experiments with added chromate substances.

Week Beginning 14/09

Summary

During this week the samples from the previous plate reader were retrieved and prepared for blotting against both GFP, and the 6His-Tag of ChrB and ChrBopt.

Day 1 - 14/09: Western blot of plate reader samples MG1655 + pSB1C3-pChr-GFP + pUniprom-ChrB, and MG1655 + pSB1C3-pChr-GFP + pUniprom-ChrBopt.

Aim of experiment: To identify whether or not there is any detectable difference in both, the level of GFP, and the level of ChrB or ChrBopt, before and after induction with IPTG.

Protocols Used: SDS-PAGE + Western blot

Results: Insert blot image here.

Next Steps: Repeat the assays of the previous weeks by subcloning ChrB, ChrBopt, and ChrB (Bielefeld) into vectors with different expression levels and see how it influences the level of GFP expression. Then add different chromate salts and also probe the system with other substances, e.g. other heavy-metal salts.

Reactants	Volume (µl)
DNA	1
DMSO	2.5
Forward Primer	0.5
Reverse Primer	0.5
Herculase Buffer	10
dNTP	0.5
Herculase	0.5
H₂O	34.5

Reactants	Volume (µl)
DNA	5
DMSO	1
Forward Primer	0.5
Reverse Primer	0.5
+Mg Buffer	2
dNTP	0.2
Taq-polymerase	0.2
H₂O	10.6

	Post Plasmid Purification	Post PCR	gBlock
DNA	1mg	50µl	100ng
Cutsmart Buffer (µl)	6	6	6
Water (µl)	As required	1	As required
Enzyme 1 (µl)	1.5	1.5	1.5
Enzyme 2 (µl)	1.5	1.5	1.5
Total Volume (µl)	30	60	30

	Vector (µl)	2:1 (Vector: Insert) [µl]	3:1 (Vector: Insert) [µl]	4:1 (Vector: Insert) [µl]
T4 DNA Ligase	1	1	1	1
Ligase Buffer	1	1	1	1
Vector	1	1	1	1
Insert	-	2	3	4
H₂0	7	5	4	3

@@ Line 74: / Line 74: @@
-          <p class="journal-content">This week was all about getting started with the newly arrived biobrick <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a></p>
+          <p class="journal-content">This week was all about getting started with the newly arrived biobrick <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008.</a></p>
           </div>
           <div id="collapsiblew1" class="collapse week-content">
@@ Line 86: / Line 86: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 22/06: Plated <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> from agar stab.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew1-1"></span>
                  <div id="collapsiblew1-1" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To prepare <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> for sequence confirmation, storage, and further processing.</p>
+                   <p><b>Aim of experiment:</b> To prepare <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> for sequence confirmation, storage, and further processing. The culture was plated from the agar stab and also a liquid culture set up.</p>
                    <p><b>Protocols Used:</b>
@@ Line 111: / Line 111: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 2</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 2 - 23/06: Purified <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> from liquid culture.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew1-2"></span>
                  <div id="collapsiblew1-2" class="collapse box-content">
@@ Line 119: / Line 119: @@
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Click here to see our miniprep protocol.</a></p>
+                     <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Miniprep plasmid purification, </a>
-                     <p><a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol"> Click here to see our sequencing protocol.</a></p>
+                     <a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol"> DNA sequencing</a>
                    </p>
                    <p><b>Results Sequencing: </b><a data-toggle="modal" data-target="#BBa_K1058008-figure1-modal" class="journal-protocol"> Figure 1 </a></p>
-                   <p><b>Next Steps:</b> PCR of parts of <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a>.</p>
+                   <p><b>Next Steps:</b> PCR of promoter region (<i>pChr</i>) and repressor region (<i>ChrB</i>) of <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> for storage in individual biobricks.</p>
@@ Line 151: / Line 151: @@
-          <p class="journal-content"> During this week, <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> was disassembled into its constituent parts, pChr, and ChrB.</p>
+          <p class="journal-content"> During this week, <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> was disassembled into its constituent parts, <i>pChr</i>, and <i>ChrB</i>.</p>
           </div>
           <div id="collapsiblew2" class="collapse week-content">
@@ Line 163: / Line 163: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 29/06: PCR-amplification of <i>pChr</i> and <i>ChrB</i> from <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a>.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew2-1"></span>
                  <div id="collapsiblew2-1" class="collapse box-content">
-                   <p><b>Aim of experiment: </b> To break down <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> into the promoter region pChr, and the repressor gene, ChrB, while adding standard prefix and suffix to each. Furthermore optimisation of first 15 codons of ChrB in order to increase the overall rate of transcription. Restriction digest of the respective ends and subsequent ligation of pChr and ChrB into pSB1C3 for submission to registry.</p>
+                   <p><b>Aim of experiment: </b> To break down <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> into the promoter region <i>pChr</i>, and the repressor gene, <i>ChrB</i>, while adding standard prefix and suffix to each. Furthermore optimisation of first 15 codons of <i>ChrB</i> in order to increase the overall rate of transcription. Restriction digest of the respective ends and subsequent ligation of <i>pChr</i> and <i>ChrB</i> into pSB1C3 for submission to registry.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> Click here to see our PCR protocol.</a></p>
+                     <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR, </a>
-                     <p><a data-toggle="modal" data-target="#gelextraction-modal" class="journal-protocol"> Click here to see our gel extraction protocol.</a></p>
+                     <a data-toggle="modal" data-target="#gelextraction-modal" class="journal-protocol"> Gel extraction, </a>
-                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol"> Click here to see our restriction digest protocol.</a></p>
+                     <a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol"> Restriction Digest</a>
                    </p>
                    <p><b>Results:</b>
-                   <p><a data-toggle="modal" data-target="#dundee15-chr-igem022-modal" class="journal-protocol"> Figure 2: Agarose gel after PCR of pChr and ChrB. </a></p>
+                   <a data-toggle="modal" data-target="#dundee15-chr-igem022-modal" class="journal-protocol"> Figure 2</a>
-                   <p>As can be seen on the image of the gel, PCR of ChrB was not successful. pChr was further processed by restriction digest with EcoRI and PstI</p></p>
+                   </p>
-                   <p><b>Next Steps:</b> Repeat of PCR of ChrB. Purification of the Plasmid, containing <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> and confirmation of size and sequence.</p>
+                   <p><b>Next Steps:</b> Repeat of PCR of <i>ChrB</i>. Purification of the Plasmid, containing <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> and confirmation of size and sequence.</p>
                  </div>
@@ Line 208: / Line 208: @@
-          <p class="journal-content">During this week ligations of pChr and ChrB into pSB1C3 were finalised and the constituent parts of our newly designed chromate sensing system were prepared for ligation and transformation.</p>
+          <p class="journal-content">During this week ligations of <i>pChr</i> and <i>ChrB</i> into pSB1C3 were finalised and the constituent parts of our newly designed chromate sensing system were prepared for ligation and transformation.</p>
           </div>
           <div id="collapsiblew3" class="collapse week-content">
@@ Line 220: / Line 220: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 06/07: Repeated amplification of <i>ChrB</i>. Ligation and transformation of <i>pChr</i> and <i>ChrB</i>.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew3-1"></span>
                  <div id="collapsiblew3-1" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To repeat PCR of ChrB for subsequent ligation into pSB1C3. Ligation of pChr and ChrB into pSB1C3. Transformation of each into a non-pathogenic lab strain of E. coli.
+                   <p><b>Aim of experiment:</b> To repeat PCR of ChrB for subsequent ligation into pSB1C3. Ligation of <i>pChr</i> and <i>ChrB</i> into pSB1C3. Transformation of each into a non-pathogenic lab strain of <i>Escherichia coli</i>.
-                   <p>Overnight culture of strain, containing GFPmut2, as a fluorescent reporter for our Chromate sensing system</p></p>
+                   Overnight culture of <i>E. coli DH5-alpha</i>, containing GFPmut2, as a fluorescent reporter for our chromate sensing system</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> Click here to see our PCR protocol.</a> <p>The annealing temperature was lowered in order to increase the chances of accomodating a primer that is not 100% complementary. Furthermore different concentrations of DMSO were tested.</p></p>
+                     <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR-amplification, </a>
-                     <p><a data-toggle="modal" data-target="#gelextraction-modal" class="journal-protocol"> Click here to see our gel extraction protocol.</a></p>
+                     <a data-toggle="modal" data-target="#gelextraction-modal" class="journal-protocol"> Gel extraction, </a>
-                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol"> Click here to see our restriction digest protocol.</a></p>
+                     <a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol"> Restriction digest, </a>
-                     <p><a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Click here to see our ligation protocol.</a> Ligations were made in 2:1 and 3:1 ratio.</p>
+                     <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligation, </a>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Click here to see our Overnight culture protocol.</a></p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight cultures</a>
                    </p>
                    <p><b>Results:</b>
-                   <p><a data-toggle="modal" data-target="#dundee15-chr-igem023-modal" class="journal-protocol"> Figure 3: Agarose gel after PCR of ChrB. </a></p>
+                   <a data-toggle="modal" data-target="#dundee15-chr-igem023-modal" class="journal-protocol"> Figure 3 </a></p>
-                   <p>2 bands were extracted from the gel for further characterisation, the strongest band, hence called ChrBs, and the band directly above, hence called ChrBw.</p>
+                   <p>2 bands were extracted from the gel for further characterisation, the strongest band, hence called <i>ChrBs</i>, and the band directly above, hence called <i>ChrBw</i>. <i>pChr</i>, <i>ChrBs</i>, and <i>ChrBw</i> were ligated into pSB1C3.
-                  <p>pChr, ChrBs, and ChrBw were ligated into pSB1C3.</p>
+                   pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw were transformed into <i>E. coli MC1061.</i></p>
-                   <p>pChr-pSB1C3, ChrBs-pSB1C3, and ChrBw-pSB1C3 were transformed into E. coli MC1061.</p></p>
-                   <p><b>Next Steps:</b> Purification of the Plasmid, containing <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> and confirmation of size and sequence.</p>
+                   <p><b>Next Steps:</b> Sequence and size confirmation of pChr-pSB1C3, ChrBs-pSB1C3, and ChrBw-pSB1C3. Ligation of <i>pChr</i> to <i>GFP coding sequence</i> for construction of a reporter plasmid.</p>
                  </div>
@@ Line 254: / Line 253: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 2</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 2 - 07/07: Purification of <i>GFP</i> and overnight cultures of pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew3-2"></span>
                  <div id="collapsiblew3-2" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> Purification of GFPmut2 from overnight cultures. Overnight cultures of recombinant MC1061, containing pChr-pSB1C3, ChrBs-pSB1C3, and ChrBw-pSB1C3 respectively. Overnight culture of pUniprom for subsequent ligation of ChrB</p>
+                   <p><b>Aim of experiment:</b> Purification of GFPmut2 from overnight cultures. Overnight cultures of recombinant <i>E. coli MC1061</i>, containing pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw respectively for further size and sequence confirmation. Overnight culture of pUniprom for subsequent ligation of ChrB</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Miniprep</a> for plasmid purification of overnight culture of GFPmut2.</p>
+                     <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Miniprep, </a>
-                     <p><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a> for amplification of GFPmut2 and ChrB.</p>
+                     <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR-amplification, </a>
-                     <p><a data-toggle="modal" data-target="#gelextraction-modal" class="journal-protocol"> Gel extraction</a> of PCR product pf GFPmut2.</p>
+                     <a data-toggle="modal" data-target="#gelextraction-modal" class="journal-protocol"> Gel extraction, </a>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight culture</a> of MC1061 containing pChr-pSB1C3, ChrBs-pSB1C3, and ChrBw-pSB1C3 respectively. Furthermore of a strain containing the vector pUniprom.</p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight cultures</a>
                    </p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-igem024-modal" class="journal-protocol"> Figure 5 </a> Since amplification of ChrB was unsuccessful, it will be repeated.</p>
+                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-igem024-modal" class="journal-protocol"> Figure 5 </a> </p>
-                   <p><b>Next Steps:</b> Sequence and size confirmation of pChr-pSB1C3, ChrBs-pSB1C3, and ChrBw-pSB1C3.</p>
+                   <p><b>Next Steps:</b> Repeat of amplification of <i>ChrB</i>. Sequence and size confirmation of pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw.</p>
                  </div>
@@ Line 283: / Line 282: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 3</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 3 - 08/07: Purification of plasmids pSB1C3-pChr, pSB1C3-ChrBs, and pSB1C3-ChrBw. Size and sequence confirmation.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew3-3"></span>
                  <div id="collapsiblew3-3" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To purify overnight cultures for size and sequence confirmation. Repeat of PCR amplification of pChr and ChrB. Restriction digestes of all parts in order to produce compatible sticky ends.</p>
+                   <p><b>Aim of experiment:</b> To purify overnight cultures for size and sequence confirmation. Repeat of PCR amplification of <i>pChr</i> and <i>ChrB</i>. Restriction digestes of all parts in order to produce compatible sticky ends.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Miniprep</a> of pChr-pSB1C3, ChrBs-pSB1C3, ChrBw-pSB1C3 and pUniprom.</p></p>
+                     <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Miniprep, </a>
-                     <p><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a> for amplification of pChr and ChrB.</p></p>
+                    <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR amplification, </a>
-                     <p><a data-toggle="modal" data-target="#restriction_digest-modal" class="journal-protocol">Restriction digest</a> for confirmation of size of pChr and ChrB.</p>
+                     <a data-toggle="modal" data-target="#restriction_digest-modal" class="journal-protocol">Restriction digest, </a>
+                    <a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a>
+                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol"> Restriction digest</a> of gel purified PCR-products of pChr, ChrB, and GFPmut2, using EcoRI/SpeI, BamHI/HindIII, and XbaI/PstI respecitvely.</p>
                    </p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-igem026-modal" class="journal-protocol"> Figure 6 </a> Agarose gel for size confirmation.</p>
+                   <p><b>Results:</b><p><a data-toggle="modal" data-target="#dundee15-chr-igem026-modal" class="journal-protocol"> Figure 6 </a> - Size confirmation of <i>pChr</i> and <i>ChrB</i>.</p>
+                   <p><a data-toggle="modal" data-target="#dundee15-chr-pChr1-modal" class="journal-protocol"> Figure 7 </a> Sequence confirmation of <i>pChr</i></p>
+                  <p><a data-toggle="modal" data-target="#dundee15-chr-ChrBw4-modal" class="journal-protocol"> Figure 8 </a> Sequence confirmation of <i>ChrB</i>.</p>
-                   <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a> of pChr and ChrB that showed expectes sizes.</p>
                    </p>
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-pChr1-modal" class="journal-protocol"> Figure 7 </a> Confirmed sequence of pChr1.</p>
+                   <p><b>Next Steps:</b> Purification and digest of pUniprom for insertion of <i>ChrB</i>.</p>
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-ChrBw4-modal" class="journal-protocol"> Figure 8 </a> Confirmed sequence of ChrBw4.</p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol"> Restriction digest</a> of gel purified PCR-products of pChr, ChrB, and GFPmut2, using EcoRI/SpeI, BamHI/HindIII, and XbaI/PstI respecitvely.</p>
-                  </p>
-                   <p><b>Next Steps:</b> Purification of the Plasmid, containing <a href="http://parts.igem.org/Part:BBa_K1058008">BBa_K1058008</a> and confirmation of size and sequence.</p>
                  </div>
@@ Line 322: / Line 313: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 4</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 4 - 09/07: Digested and cleaned up pUniprom for insertion of <i>ChrB</i>. Amplification of optimised <i>ChrB</i> with primers specific for the insertion into pUniprom.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew3-4"></span>
                  <div id="collapsiblew3-4" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To prepare pUniprom for insertion of ChrB.</p>
+                   <p><b>Aim of experiment:</b> To prepare pUniprom for insertion of <i>ChrB</i> by digesting it with BamHI and HindIII. To amplify <i>ChrB</i> version with first 15 codons optimised for insertion into pUniprom.</p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> of pUniprom with BamHI and HindIII.</p>
-                    <p><a data-toggle="modal" data-target="#gel-extraction-modal" class="journal-protocol">Gel extraction</a> of digested pUniprom</p>
-                  </p>
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-igem034-modal" class="journal-protocol"> Figure 9 </a></p>
-                  <p><b>Next Steps:</b> Ligation of ChrB into pUniprom.</p>
-                  <p><b>Aim of experiment:</b> To amplify ChrB version with first 15 codons optimised for insertion into pUniprom.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR</a> of optimised ChrB from the sequence confirmed ChrBw4-pSB1C3, adding restriction sites compatible to pUniprom.</p>
+                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> and
-                     <p><a data-toggle="modal" data-target="#gelextraction" class="journal-protocol">Gel extraction</a> of optimised ChrBafter PCR.</p>
+                    <a data-toggle="modal" data-target="#gel-extraction-modal" class="journal-protocol">Gel extraction</a> of pUniprom</p>
+                     <p><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol">PCR</a> and
+                    <a data-toggle="modal" data-target="#gelextraction" class="journal-protocol"> Gel extraction </a> of optimised <i>ChrB</i></p>
                    </p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-igem034-modal" class="journal-protocol"> Figure 10 </a></p>
+                   <p><b>Results:</b><p><a data-toggle="modal" data-target="#dundee15-chr-igem034-modal" class="journal-protocol"> Figure 9 </a> pUniprom</p>
+                  <p><a data-toggle="modal" data-target="#dundee15-chr-igem032-modal" class="journal-protocol"> Figure 10 </a> ChrB</p></p>
-                   <p><b>Next Steps:</b> Ligation of ChrB and codon optimised ChrB into pUniprom.</p>
+                   <p><b>Next Steps:</b> Ligation of <i>ChrB</i> and codon optimised <i>ChrB</i> into pUniprom.</p>
                  </div>
@@ Line 361: / Line 342: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 5</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 5 - 10/07: Ligations of <i>ChrB</i> and optimised <i>ChrB</i> into pUniprom, and of <i>pChr</i> and <i>GFPmut2</i> into pSB1C3.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew3-5"></span>
                  <div id="collapsiblew3-5" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> Ligation of ChrB and optimised ChrB into pUniprom. Ligation of pChr and GFPmut2 into pSB1C3</p>
+                   <p><b>Aim of experiment:</b> Ligation of <i>ChrB</i> and optimised <i>ChrB</i> into pUniprom via BamHI and HindIII restriction sites. Ligation of <i>pChr</i> and <i>GFPmut2</i> to each other via SpeI/XbaI-link and into pSB1C3 via EcoRI and PstI restriction sites.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation</a> of ChrB and optimised ChrB into pUniprom via BamHI and HindIII restriction sites. Ligation of pChr to GFPmut2 via Spe/Xba link and ligation of pChrGFP construct into pSB1C3 EcoRI/PstI restriction sites.</p>
+                     <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligations</a>
                    </p>
                    <p><b>Results:</b> N/A</p>
-                   <p><b>Next Steps:</b>Transformation of abovementioned ligations.</p>
+                   <p><b>Next Steps:</b> Transformation of abovementioned ligations.</p>
@@ Line 401: / Line 382: @@
-          <p class="journal-content">During this week successful transformations were identified and propagated.</p>
+          <p class="journal-content">During this week successful transformants were identified and propagated.</p>
           </div>
           <div id="collapsiblew4" class="collapse week-content">
@@ Line 413: / Line 394: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 13/07: Transformation of pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP.</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew4-1"></span>
                  <div id="collapsiblew4-1" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> Transform recombinant vectors into E. coli chassis.</p>
+                   <p><b>Aim of experiment:</b> Transformation of pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP into <i>E. coli JM110</i> chassis. Setting up a backup restriction digest of pSB1C3 with EcoRI and SpeI in case the double ligation pSB1C3-pChr-GFP is unsuccessful.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> of ChrB-pUniprom, optimised ChrB-pUniprom and pChr-GFPmut2-pSB1C3 into JM110.</p>
+                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformations, </a>
+                    <a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a>
                    </p>
                    <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-first-transformation-modal" class="journal-protocol"> Figure 11 </a></p>
-                   <p><b>Aim of experiment:</b> Backup of digested pSB1C3 for the case that a ligation with 2 inserts does not work.</p>
+                   <p><b>Next Steps:</b> Purification of recombinant vectors and confirmation of size and sequence. Store digested pSB1C3 at -20<sup>o</sup>C.</p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> of pSB1C3 with EcoRI/SpeI for subsequent insertion of pChr.</p>
-                  </p>
-                  <p><b>Results:</b>N/A</p>
-                  <p><b>Next Steps:</b> Store digested plasmid at -20<sup>o</sup>C.</p>
                  </div>
-        </div>
+          </div>
          </div>
        </div>
@@ Line 446: / Line 419: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 2</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 2 - 14/07: Set up cultures for subsequent purification of recombinant vectors for size and sequence confirmation.</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew4-2"></span>
                  <div id="collapsiblew4-2" class="collapse box-content">
+                   <p><b>Aim of experiment:</b> Purification of recombinant vectors, pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP, for size and sequence confirmation. Patch plating of the same colonies for storage and retrieval once size and sequence have been confirmed.</p>
-                   <p><b>Aim of experiment:</b> Purification of recombinant vectors for sequence and size confirmation.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight culture</a> of JM110 with ChrB-pUniprom, optimised ChrB-pUniprom and pChr-GFPmut2-pSB1C3.</p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight cultures, </a>
-                   <p><a data-toggle="modal" data-target="#plating-modal" class="journal-protocol"> Patch plating</a> of the same colonies.</p>
+                    <a data-toggle="modal" data-target="#plating-modal" class="journal-protocol"> Patch plating</a>
                    </p>
@@ Line 472: / Line 444: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 3</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 3 - 15/07: Purified pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP from overnight cultures</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew4-3"></span>
                  <div id="collapsiblew4-3" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> Purification of recombinant plasmids for confirmation of size and sequence.</p>
+                   <p><b>Aim of experiment:</b> Purification of pUniprom-ChrB, pUniprom-ChrBopt, and pSB1C3-pChr-GFP from overnight for confirmation of size and sequence. Restriction digest of pUnipromChrB and optimised pUniprom-ChrB optimised with BamHI and HindIII and of pSB1C3-pChr-GFP with EcoRI/PstI for size confirmation. </p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Miniprep</a> of ChrB-pUniprom, optimised ChrB-pUniprom and pChr-GFPmut2-pSB1C3.</p>
+                     <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Miniprep, </a>
+                    <a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest, </a>
+                    <a data-toggle="modal" data-target="#colony-modal" class="journal-protocol">Colony PCR, </a>
+                    <a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing </a>
                    </p>
-                   <p><b>Results:</b>N/A</p>
+                   <p><b>Results:</b>
+                    <p><a data-toggle="modal" data-target="#dundee15-chr-igem036-modal" class="journal-protocol"> Figure 12 </a> Size confirmation of <i>ChrB</i> and <i>ChrBopt</i>.</p>
-                 <p><b>Protocols Used:</b>
+                    <a data-toggle="modal" data-target="#dundee15-chr-igem037-modal" class="journal-protocol"> Figure 13 </a> Gel after colony PCR. Colonies 2, 4, 9, and 12 were selected for sequencing.</p>
-                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> of ChrB-pUniprom and optimised ChrB-pUniprom with BamHI and HindIII. Restriction digest of pChr-GFPmut2-pSB1C3 with EcoRI/PstI.</p>
+                     </p><a data-toggle="modal" data-target="#dundee15_chr_pSB1C3-pChr-GFP_reverse" class="journal-protocol"> Figure 14 </a> Sequencing result of pSB1C3-pChr-GFPmut2_1. The Result for pSB1C3-pChr-GFPmut2_2 was not as expected, albeit in the correct orientation.</p>
                    </p>
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-igem036-modal" class="journal-protocol"> Figure 1 </a></p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#colony-modal" class="journal-protocol">Colony PCR</a> of pUniprom-ChrB-opt for elucidation of insert size or presence of insert.</p>
-                  </p>
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15-chr-igem037-modal" class="journal-protocol"> Figure 13 </a> Gel after colony PCR. Colonies 2, 4, 9, and 12 were selected for sequencing.</p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a> of pSB1C3-pChr-GFP.</p>
-                  </p>
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_pSB1C3-pChr-GFP_reverse" class="journal-protocol"> Figure 14 </a> Sequencing result of pSB1C3-pChr-GFPmut2_1. The Result for pSB1C3-pChr-GFPmut2_2 was not as expected, albeit in the correct orientation.</p>
-                   <p><b>Next Steps:</b> Confirmation of insert size and sequence of ChrB-opt.</p>
+                   <p><b>Next Steps:</b> Confirmation of sequence of <i>ChrB-opt.</i></p>
@@ Line 515: / Line 476: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 4</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 4 - 16/07: Set up overnight cultures of colonies chosen after colony-PCR for subsequent purification and sequence confirmation.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew4-4"></span>
                  <div id="collapsiblew4-4" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To prepare ChrB-opt colonies 2, 4, 9, and 12 for sequencing.</p>
+                   <p><b>Aim of experiment:</b> Set up overnight cultures of <i>ChrBopt</i> colonies 2, 4, 9, and 12 for sequencing.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Click here to see our overnight culture protocol.</a></p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight cultures </a></p>
                    </p>
@@ Line 540: / Line 501: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 5</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 5 - 17/07: Purified and sequenced chosen <i>ChrBopt</i>-colonies.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew4-5"></span>
                  <div id="collapsiblew4-5" class="collapse box-content">
@@ Line 548: / Line 509: @@
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Miniprep</a> of ChrB-opt 2, 4, 9, 12.</p>
+                     <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Miniprep, </a>
+                    <a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a>
                    </p>
-                   <p><b>Results:</b>N/A</p>
+                   <p><b>Results:</b> <i>ChrBopt</i> showed HindIII restriction sites and was hence cleaved into partial inserts.</p>
-                   <p><b>Protocols Used:</b>
+                   <p><b>Next Steps:</b> Use alternative cloning protocol for <i>ChrB</i> and <i>ChrBopt</i>.</p>
-                    <p><a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a> of ChrB-opt 2, 4, 9, 12.</p>
-                  </p>
-                  <p><b>Results:</b> ChrB showed HindIII restriction sites and was hence cleaved into partial inserts.</p>
-                  <p><b>Next Steps:</b>Use alternative cloning protocol for ChrB and ChrBopt.</p>
                  </div>
-        </div>
+          </div>
          </div>
        </div>
@@ Line 586: / Line 542: @@
-          <p class="journal-content">During this week an alternative cloning strategy for ChrB and ChrB-opt was employed. Furthermore, the pChr-GFP construct was cloned in a stepwise fashion.</p>
+          <p class="journal-content">During this week an alternative cloning strategy for <i>ChrB</i> and <i>ChrBopt</i> was employed. Furthermore, the pSB1C3-pChr-GFP construct was cloned in a stepwise fashion.</p>
           </div>
           <div id="collapsiblew5" class="collapse week-content">
@@ Line 598: / Line 554: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1 - 20/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 20/07: Went back to the original plate of pSB1C3-pChr-GFP for visualising colonies that do or do not express GFP. </span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew5-1"></span>
                  <div id="collapsiblew5-1" class="collapse box-content">
+                   <p><b>Aim of experiment:</b>To compare colonies on the patch plate of pSB1C3-pChr-GFP by differentially visualising colonies that do or do not express GFP under UV light. The background of this experiment was that if there is one colony that has both inserts in the correct orientation with respect to each other, then there is perhaps one that also has them with the correct orientation with respect to the plasmid. Colonies identified as expressing GFP were patch plated and grown in liquid cultures over night in order to confirm the sequence of the insert.</p>
-                   <p><b>Aim of experiment:</b>To compare colonies on the patch plate of pSB1C3-pChr-GFP by differentially visualising colonies that do or do not express GFP under UV light.</p>
-                  <!--
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#plating-modal" class="journal-protocol"> Click here to see our plating protocol.</a></p>
+                     <a data-toggle="modal" data-target="#plating-modal" class="journal-protocol"> Patch-plating, </a>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Click here to see our overnight culture protocol.</a></p></p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> overnight cultures</a>
+                   </p>
-                   <p><b>Results:</b> N/A</p>
-                  -->
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_pChrGFPplates_revisited" class="journal-protocol">Figure 15</a> pSB1C3-pChr-GFP in JM110.</p>
+                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_pChrGFPplates_revisited" class="journal-protocol">Figure 15</a> pSB1C3-pChr-GFP in <i>E. coli JM110</i>.</p>
-                   <p><b>Next Steps:</b>Size confirmation and sequencing of newly selected colonies.</p>
+                   <p><b>Next Steps:</b> Sequence confirmation of newly selected colonies.</p>
                  </div>
-        </div>
+          </div>
          </div>
        </div>
@@ Line 629: / Line 579: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 2 - 21/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 2 - 21/07: Purified plasmids from new colonies and confirmed sequence. Restriction digest of pUniprom with BamHI and PstI.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew5-2"></span>
                  <div id="collapsiblew5-2" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>Purification of newly selected colonies containing pSB1C3-pChr-GFP for subsequent sequence confirmation.</p>
+                   <p><b>Aim of experiment:</b>Purification of newly selected colonies, 1, 5, 6, and 7, containing pSB1C3-pChr-GFP for subsequent sequence confirmation. Restriction digest of pUniprom with BamHI and PstI to employ an alternative cloning strategy of <i>ChrB</i> and <i>ChrBopt</i>.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Miniprep</a> for purification of pSB1C3-pChr-GFP.</p>
+                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Miniprep </a> and
+                    <a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing </a> of pChrGFP insert.</p>
+                    <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> of pUniprom.
                    </p>
-                   <p><b>Results:</b>N/A</p>
+                   <p><b>Results:</b>
+                  <p><a data-toggle="modal" data-target="#dundee15_chr_pChrGFP_revisited_seq" class="journal-protocol"> Figure 16 </a> Alignment of sequencing results.</p>
+                  <p><a data-toggle="modal" data-target="#dundee15_chr_igem039" class="journal-protocol"> Figure 17 </a> Gel after restriction digest of pUniprom.</p>
-                   <p><b>Next Steps:</b>Sequencing of pSB1C3-pChr-GFP samples 1, 5, 6, and 7.</p>
+                   <p><b>Next Steps:</b>Stepwise ligation of <i>pChr</i> and <i>GFP</i> into pSB1C3 in order to clone the inserts in the correct orientation with respect to the plasmid. Insertion of <i>ChrB</i> and <i>ChrBopt</i> into pUniprom via BamHI and PstI links.</p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a> of pSB1C3-pChr-GFP samples 1, 5, 6, and 7.</p>
-                  </p>
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_pChrGFP_revisited_seq" class="journal-protocol"> Figure 16 </a> Alignment of sequencing results.</p>
-                  <p><b>Next Steps:</b>Stepwise ligation of pChr-GFP into pSB1C3.</p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> of pUniprom with BamHI/PstI for subsequent insertion of ChrB and ChrB-opt.</p>
-                  </p>
-                  p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_igem039" class="journal-protocol"> Figure 17 </a> Gel image</p>
-                  <p><b>Next Steps:</b>Insertion of ChrB and ChrB-opt respectively via BamHI/PstI.</p>
                  </div>
-        </div>
+          </div>
          </div>
        </div>
@@ Line 670: / Line 606: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 3 - 22/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 3 - 22/07: Ligated <i>pChr</i> into pSB1C3 via EcoRI and SpeI sites and cloned the recombinant plasmid into <i>E. coli JM110</i>.</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew5-3"></span>
                  <div id="collapsiblew5-3" class="collapse box-content">
+                   <p><b>Aim of experiment:</b>Ligation of <i>pChr</i> into pSB1C3 via EcoRI SpeI link. This is the first step of a 2-step cloning strategy with the aim of cloning <i>pChr</i> and <i>GFP</i> into pSB1C3 in the correct orientation with respect to the plasmid.</p>
-                   <p><b>Aim of experiment:</b>Ligation of pChr into pSB1C3 via Eco/Spe link.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation</a> of pChr(08/07/15) into pSB1C3(13/07/15)</p>
+                     <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation, </a>
-                  </p>
+                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a>
-                  <p><b>Results:</b> N/A</p>
-                  <p><b>Aim of experiment:</b>Transformation</p>
-                  <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> of pSB1C3-pChr into JM110.</p>
                    </p>
                    <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150723_pSB1C3-pChr" class="journal-protocol"> Figure 18 </a> Plates after transformation of pChr into pSB1C3.</p>
-                   <p><b>Next Steps:</b>Sequence confirmation of pChr in pSB1C3.</p>
+                   <p><b>Next Steps:</b> Sequence confirmation of pChr in pSB1C3.</p>
                  </div>
-        </div>
+         </div>
          </div>
        </div>
@@ Line 703: / Line 630: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 4 - 23/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 4 - 23/07: Set up overnight cultures of pSB1C3-pChr colonies.</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew1-4"></span>
                  <div id="collapsiblew1-4" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To prepare pSB1C3-pChr for sequence confirmation</p>
+                   <p><b>Aim of experiment:</b> Set up overnight cultures of pSB1C3-pChr colonies to prepare pSB1C3-pChr for sequence confirmation</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol">Overnight culture</a> of pSB1C3-pChr.</p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol">Overnight culture, </a>
-                     <p><a data-toggle="modal" data-target="#plating-modal" class="journal-protocol">Patch-plating</a> of the same colonies of pSB1C3-pChr.</p>
+                     <a data-toggle="modal" data-target="#plating-modal" class="journal-protocol">Patch-plating</a>
                    </p>
                    <p><b>Results:</b> N/A</p>
-                   <p><b>Next Steps:</b> Purification of the recombinant vector for sequence confirmation.</p>
+                   <p><b>Next Steps:</b> Purification of the recombinant vector for size and sequence confirmation.</p>
                  </div>
-        </div>
+          </div>
          </div>
        </div>
@@ Line 729: / Line 656: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 5 - 24/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 5 - 24/07: Purified pSB1C3-pChr for sequence confirmation.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew5-5"></span>
                  <div id="collapsiblew5-5" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>Purification of pSB1C3-pChr for sequence confirmation.</p>
+                   <p><b>Aim of experiment:</b> Purification of pSB1C3-pChr for sequence confirmation before ligating <i>GFP</i> into it.</p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Miniprep</a> of pSB1C3-pChr.</p>
-                  </p>
-                  <p><b>Results:</b> Insert table here</p>
-                  <p><b>Aim of experiment:</b>Sequence confirmation of pSB1C3-pChr.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a> of samples 1 and 2 of pSB1C3-pChr.</p>
+                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol"> Plasmid purification, </a>
+                    <a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing </a>
                    </p>
@@ Line 763: / Line 682: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 6 - 25/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 6 - 25/07: PCR-amplified <i>ChrB</i> and <i>ChrBopt</i>.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew5-6"></span>
                  <div id="collapsiblew5-6" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> Amplification of ChrB and optimised ChrB</p>
+                   <p><b>Aim of experiment:</b> Amplification of <i>ChrB</i> and <i>ChrBopt</i> for insertion into pUniprom. Restriction digest of the PCR product with BamHI and PstI for producing compatible sticky ends</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR</a>-mplification of ChrB and ChrBopt.</p>
+                     <a data-toggle="modal" data-target="#PCR-modal" class="journal-protocol"> PCR-amplification, </a>
+                    <a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol"> Restriction digest</a>
                    </p>
                    <p><b>Results:</b> N/A</p>
-                   <p><b>Next Steps:</b>Ligation and Transformation of ChrB and optimised ChrB.</p>
+                   <p><b>Next Steps:</b>Ligation and Transformation of <i>ChrB</i> and <i>ChrBopt</i>.</p>
                  </div>
-        </div>
+          </div>
          </div>
        </div>
@@ Line 788: / Line 708: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 7 - 26/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 7 - 26/07: Ligated <i>ChrB</i> and <i>ChrBopt</i> into pUniprom.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew5-7"></span>
                  <div id="collapsiblew5-7" class="collapse box-content">
@@ Line 796: / Line 716: @@
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligation</a> of ChrB and optimised ChrB into vector pUniprom.</p>
+                     <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol"> Ligation</a>
                    </p>
@@ Line 840: / Line 760: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1 - 27/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 27/07: Transformed <i>ChrB</i> and <i>ChrBopt</i> into <i>E. coli JM110</i></span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew6-1"></span>
                  <div id="collapsiblew6-1" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>Transformation of the repressors, ChrB and optimised ChrB, into JM110</p>
+                   <p><b>Aim of experiment:</b>Transformation of the repressors, <i>ChrB</i> and <i>ChrBopt</i>, into <i>JM110</i></p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> of ChrB and optimised ChrB into JM110</p>
+                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a>
                    </p>
+                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150728_pUniprom-ChrB" class="journal-protocol"> Figure 20</a> Plates after transformation.</p>
-                  <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150728_pUniprom-ChrB" class="journal-protocol"> Figure 1 </a></p>
+                   <p><b>Next Steps:</b> Purification of pUniprom-ChrB and pUniprom-ChrBopt for sequence confirmation.</p>
-                   <p><b>Next Steps:</b> Purification of the plasmids for sequence confirmation.</p>
                  </div>
-        </div>
+         </div>
          </div>
        </div>
@@ Line 866: / Line 785: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 2 - 28/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 2 - 28/07: Digested pSB1C3-pChr for insertion of <i>GFP</i></span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew6-2"></span>
                  <div id="collapsiblew6-2" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>Processing of pSB1C3-pChr from 24/07 for insertion of GFP.</p>
+                   <p><b>Aim of experiment:</b>Digesting sequence confirmed pSB1C3-pChr with SpeI and PstI for subsequent ligation with <i>GFP</i>. Overnight cultures of <i>ChrB</i> and <i>ChrBopt</i> for subsequent plasmid purification, size confirmation, and sequence confirmation.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> of pSB1C3-pChr with SpeI/PstI for insertion of GFP.</p>
+                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest, </a>
-                     <p><a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation</a> of GFPmut2, previously digested with XbaI/PstI.</p>
+                    <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation</a> of <i>GFP</i> into pSB1C3-pChr.</p>
+                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol">Overnight culture</a> of of <i>ChrB</i> and <i>ChrBopt</i>.</p>
                    </p>
-                  <p><b>Results:</b> N/A</p>
                    <p><b>Next Steps:</b> Transformation of recombinant vector for confirmation of size and sequence.</p>
-                  <p><b>Aim of experiment:</b>Overnight culture from plates of ChrB and optimised ChrB.</p>
-                  <p><b>Protocols Used:</b>
-                    <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol">Overnight culture</a> of of ChrB and optimised ChrB.</p>
-                  </p>
                    <p><b>Results:</b> N/A</p>
-                   <p><b>Next Steps:</b> Purification of plasmids for sequence confirmation.</p>
+                   <p><b>Next Steps:</b>Transformation of pSB1C3-pChr-GFP for size and sequence confirmation. Purification of <i>ChrB</i> and <i>ChrBopt</i> for sequence confirmation.</p>
@@ Line 902: / Line 814: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 3 - 29/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 3 - 29/07: Purified <i>ChrB</i> and <i>ChrBopt</i> for size and sequence confirmation.</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew6-3"></span>
                  <div id="collapsiblew6-3" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>Purification of ChrB and optimised ChrB for confirmation of size and sequence.</p>
+                   <p><b>Aim of experiment:</b> Purification of <i>ChrB</i> and <i>ChrBopt</i> for confirmation of size and sequence.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Miniprep</a> of ChrB and ChrBopt.</p>
+                     <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid purification, </a>
-                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction Digest</a> of the purified vectors for confirmation of size.</p>
+                     <a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest, </a>
-                     <p><a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a> of ChrB and ChrBopt.</p>
+                     <a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a>
                    </p>
                    <p><b>Results:</b>
-                     <p><a data-toggle="modal" data-target="#dundee15_chr_igem040" class="journal-protocol">Confirmation of insert size.</a></p>
+                     <p><a data-toggle="modal" data-target="#dundee15_chr_igem040" class="journal-protocol">Figure 21 </a>Confirmation of insert size.</p>
-                     <p><a data-toggle="modal" data-target="#dundee15_chr_150729_chrb-combined" class="journal-protocol">Confirmation of sequence.</a></p>
+                     <p><a data-toggle="modal" data-target="#dundee15_chr_150729_chrb-combined" class="journal-protocol"> Figure 22 </a>Confirmation of sequence.</p>
                    </p>
@@ Line 932: / Line 844: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 4 - 30/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 4 - 30/07: Repeated transformation of pSB1C3-pChr-GFP into <i>E.coli JM110</i></span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew6-4"></span>
                  <div id="collapsiblew6-4" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>Digest of pSB1C3-pChr with SpeI/PstI for insertion of GFP. Repeat of unsuccessful transformation from 28/07.</p>
+                   <p><b>Aim of experiment:</b>Digest of pSB1C3-pChr with SpeI and PstI for insertion of GFP. Repeat of unsuccessful transformation from 28/07.</p>
-                   <p><b>Protocols Used:</b>
+                   <p><b>Protocols Used: </b>
-                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> of pSB1C3-pChr with SpeI/PstI for insertion of GFP.</p>
+                     <a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest, </a>
-                     <p><a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation</a> of GFPmut2, previously digested with XbaI/PstI.</p>
+                     <a data-toggle="modal" data-target="#ligation-modal" class="journal-protocol">Ligation, </a>
-                     <p><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a> of pChrGFP into JM110.</p>
+                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a>
                    </p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150731_pChrGFP-ligations" class="journal-protocol"> Figure 22 </a></p>
+                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150731_pChrGFP-ligations" class="journal-protocol"> Figure 23 </a>Plates after transformation</p>
-                   <p><b>Next Steps:</b> Purification of the Plasmids for confirmation of insert size and sequence.</p>
+                   <p><b>Next Steps:</b> Overnight cultures for purification of the pSB1C3-pChr-GFP for confirmation of insert size and sequence.</p>
@@ Line 959: / Line 871: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 5 - 31/07</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 5 - 31/07: Set up overnight cultures for confirmation of pSB1C3-pChr-GFP.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew6-5"></span>
                  <div id="collapsiblew6-5" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>Overnight cultures for subsequent plasmid purification of pSB1C3-pChr-GFP.</p>
+                   <p><b>Aim of experiment:</b> Overnight cultures for subsequent plasmid purification of pSB1C3-pChr-GFP.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol">Overnight culture</a>of pSB1C3-pChr-GFP.</p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol">Overnight culture</a>
                    </p>
@@ Line 984: / Line 896: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 6 - 01/08</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 6 - 01/08: Purified pSB1C3-pChr-GFP for size confirmation and sequencing.</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew6-6"></span>
                  <div id="collapsiblew6-6" class="collapse box-content">
@@ Line 992: / Line 904: @@
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Miniprep</a> for purification of pSB1C3-pChr-GFP.</p>
+                     <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid purification, </a>
-                     <p><a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest</a> of purified plasmid with EcoRI/PstI for confirmation of insert size.</p>
+                     <a data-toggle="modal" data-target="#restriction-digest-modal" class="journal-protocol">Restriction digest, </a>
+                    <a data-toggle="modal" data-target="#sequencing-modal" class="journal-protocol">Sequencing</a>
                    </p>
                    <p><b>Results:</b>
                      <p><a data-toggle="modal" data-target="#dundee15_chr_igem043" class="journal-protocol"> Figure 24 </a> Gel for size confirmation.</p>
-                     <p><a data-toggle="modal" data-target="#dundee15_chr_150801_pchrgfp" class="journal-protocol"> Figure 24 </a> Sequence confirmation.</p>
+                     <p><a data-toggle="modal" data-target="#dundee15_chr_150801_pchrgfp" class="journal-protocol"> Figure 25 </a> Sequence confirmation.</p>
                    </p>
-                   <p><b>Next Steps:</b>Overnight cultures in preparation for western blotting for characterisation of expression of both, GFP, and ChrB.</p>
+                   <p><b>Next Steps:</b>Overnight cultures in preparation for western blotting for characterisation of expression of both, <i>GFP</i>, and <i>ChrB</i> and <i>ChrBopt</i>.</p>
@@ Line 1,013: / Line 926: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 7 - 02/08</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 7 - 02/08: Set up cultures in preparation for western blotting.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew6-7"></span>
                  <div id="collapsiblew6-7" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>Overnight cultures of sequence confirmed ChrB, ChrBopt, pChrGFP1 (from 15/07), pChrGFP3 and pChrGFP4 (31/07) for subsequent western blotting.</p>
+                   <p><b>Aim of experiment:</b>Overnight cultures of sequence confirmed <i>ChrB</i>, <i>ChrBopt</i>, <i>pChrGFP1</i> (from 15/07), <i>pChrGFP3</i> and <i>pChrGFP4</i> (31/07) for subsequent western blotting.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight culture</a> of named constituents.</p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight culture</a>
                    </p>
                    <p><b>Results:</b> N/A</p>
-                   <p><b>Next Steps:</b> Western blotting for characterisation of expression of both, GFP, and ChrB.</p>
+                   <p><b>Next Steps:</b> Western blotting for characterisation of expression of both, <i>GFP</i>, and <i>ChrB</i> and <i>ChrBopt</i>.</p>
@@ Line 1,053: / Line 966: @@
-          <p class="journal-content">This week was entirely used for characterising the expression of GFP from the promoter pChr, and the expression of ChrB from the <a href="http://parts.igem.org/Part:BBa_K895000" target="_blank">tat-promoter</a> in pUniprom.</p>
+          <p class="journal-content">This week was entirely used for characterising the expression of GFP from the promoter <i>pChr</i>, and the expression of <i>ChrB</i> from the <a href="http://parts.igem.org/Part:BBa_K895000" target="_blank"><i>tat-promoter</i></a> in pUniprom.</p>
           </div>
           <div id="collapsiblew7" class="collapse week-content">
@@ Line 1,065: / Line 978: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1 - Day 7</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - Day 7 - 03/08 - 09/08: Determined expression of <i>ChrB</i>, <i>ChrBopt</i>, and GFP by western blotting.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew7-1"></span>
                  <div id="collapsiblew7-1" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>To determine whether or not the required proteins, GFP, and ChrB, are expressed.</p>
+                   <p><b>Aim of experiment:</b>To determine whether or not the required proteins, <i>GFP</i>, and <i>ChrB</i>, are expressed by blotting against GFP and against a 6His-tag respectively.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#sds-modal" class="journal-protocol">SDS-PAGE and western blot</a> against GFP and against His.</p>
+                     <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol">SDS-PAGE and western blot</a>
                    </p>
@@ Line 1,085: / Line 998: @@
                    <p><a data-toggle="modal" data-target="#dundee15_chr_150809_blot" class="journal-protocol"> Figure 26 </a></p>
-                   <p><b>Next Steps:</b>Transformation of both plasmids into MG1655 with the goal to characterise the interaction between ChrB and pChr. Furthermore the original biobrick, <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> will be tested under the same conditions for comparison.</p>
+                   <p><b>Next Steps:</b>Transformation of both plasmids into MG1655 with the goal to characterise the interaction between <i>ChrB</i> and <i>pChr</i>. Furthermore the original biobrick, <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> will be tested under the same conditions for comparison.</p>
@@ Line 1,113: / Line 1,026: @@
-          <p class="journal-content">This week was spent with transforming all combinations of plasmids into MG1655. Tests for their interaction were also prepared.</p>
+          <p class="journal-content">This week was spent with transforming all combinations of pSB1C3-pChr-GFP and pUniprom-ChrB into <i>E. coli MG1655</i>. Tests for their interaction were also prepared.</p>
           </div>
           <div id="collapsiblew8" class="collapse week-content">
@@ Line 1,125: / Line 1,038: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1 - 10/08</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 10/08: Transformed both pSB1C3-pChr-GFP-plasmids, pUniprom-ChrB, and pUniprom-ChrBopt, as well as <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> into <i>E. coli MG1655</i>.</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew8-1"></span>
                  <div id="collapsiblew8-1" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>To transform the following combinations of plasmids into MG1655:
+                   <p><b>Aim of experiment:</b> To transform the following combinations of pSB1C3-pChr-GFP and pUniprom-ChrB into <i>MG1655</i>:
                      <ul>
                        <li>ChrB2 (29/07) - pChrGFP (31/07)</li>
@@ Line 1,137: / Line 1,050: @@
                        <li>ChrBopt3 (29/07) - pChrGFP (15/07)</li>
                      </ul>
-                   in a stepwise fashion.</p>
+                   in a stepwise fashion. Furthermore transform <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> into MG1655. Transformations were done in a stepwise fashion with only one of the plasmids at a time.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Miniprep</a> all 4 constituents. Also purify <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a></p>
+                     <a data-toggle="modal" data-target="#miniprep-modal" class="journal-protocol">Plasmid purification, </a>
-                     <p><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transform</a> all 4 in separate aliquots of MG1655.</p>
+                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformation</a>
                    </p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150811_transformations" class="journal-protocol"> Figure 1 </a></p>
+                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150811_transformations" class="journal-protocol"> Figure 27 </a></p>
                    <p><b>Next Steps:</b>Making newly transformed cells competent again in order to transform the second plasmid as well.</p>
@@ Line 1,159: / Line 1,072: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 2 - 11/08</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 2 - 11/08: Set up overnight cultures of all transformed cells.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew8-2"></span>
                  <div id="collapsiblew8-2" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To prepare overnight cultures of the transformations in order to make them competent again.</p>
+                   <p><b>Aim of experiment:</b> To prepare overnight cultures of yesterdays transformaions in order to make them competent to transform the second plasmid as well.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol">Overnight culture</a> of a colony of each plate.</p>
+                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol">Overnight culture</a>
                    </p>
@@ Line 1,185: / Line 1,098: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 3 - 12/08: Made competent MG1655 with ChrB2, ChrBopt3, pChrGFP (15/07), pChrGFP (31/07).</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 3 - 12/08: Made competent <i>MG1655</i> with pUniprom-ChrB2, pUniprom-ChrBopt3, pSB1C3-pChr-GFP (15/07), pSB1C3-pChr-GFP (31/07).</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew8-3"></span>
                  <div id="collapsiblew8-3" class="collapse box-content">
-                   <p><b>Aim of experiment:</b>To make transformed cells competent in order to transform the second plasmid for a full system.</p>
+                   <p><b>Aim of experiment:</b>To make transformed cells competent in order to transform the second plasmid. The same plasmids that were used for the initial transformations were also used for this set of transformations.</p>
                    <p><b>Protocols Used:</b>
-                     <p><a data-toggle="modal" data-target="#competent-modal" class="journal-protocol">Making competent cells</a> of MG1655, containing ChrB2, ChrBopt3, pChrGFPrev, pChrGFPfor respectively.</p>
+                     <a data-toggle="modal" data-target="#competent-modal" class="journal-protocol">Competent cells, </a>
-                     <p><a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transform</a> same plasmids that were used for initial transformation into newly made competent cells in order to get all 4 combinations of plasmids as outline on day 1 of this week.</p>
+                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol">Transformations </a>
                    </p>
-                   <p><b>Results:</b> insert images of plates once they are round.</p>
+                   <p><b>Results:</b> N/A</p>
-                   <p><b>Next Steps:</b>Blot against GFP in order to confirm (or refute) that ChrB interacts with pChr and hence switches off expression of GFP.</p>
+                   <p><b>Next Steps:</b>Blot against GFP in order to confirm (or refute) that <i>ChrB</i> interacts with <i>pChr</i> and hence switches off expression of GFP.</p>
@@ Line 1,212: / Line 1,125: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 4 - 13/08: Transformed ChrB2 and ChrBopt3 into MG1655+pChrGFP(15/07) and MG1655+pChrGFP(31/07)</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 4 - 13/08: Transformed pUniprom-ChrB2 and pUniprom-ChrBopt3 into <i>MG1655+pSB1C3-pChr-GFP</i>(15/07) and <i>MG1655+pChrGFP</i>(31/07)</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew8-4"></span>
                  <div id="collapsiblew8-4" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To build a strain of MG1655, containing both plasmids required for a chromate sensor.</p>
+                   <p><b>Aim of experiment:</b> To build a strain of <i>E. coli MG1655</i>, containing both plasmids required for a chromate sensor.</p>
                    <p><b>Protocols Used:</b>
@@ Line 1,225: / Line 1,138: @@
                    <p><b>Results:</b> No successful transformants were found.</p>
-                  <!-- For images in the results use the following line of code:
+                   <p><b>Next Steps:</b> Repeat this experiment but transform pSB1C3-pChr-GFP (15/07) and pSB1C3-pChr-GFP (31/07) into competent <i>MG1655+pUniprom-ChrB2</i> and <i>MG1655+pUniprom-ChrBopt3</i> respectively.</p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#lbpa-figure1-modal" class="journal-protocol"> Figure 1 </a></p>
-                  under data-target refer to a modal specifically made for each image under image modals section-->
-                  <p><b>Next Steps:</b> Check Plates for successful transformants.</p>
                  </div>
@@ Line 1,256: / Line 1,162: @@
-          <p class="journal-content">During this week recombinant MG1655 were made, containing both plasmids. Western Blots were used to determine expression of ChrB, ChrBopt, and GFP.</a></p>
+          <p class="journal-content">During this week <i>E. coli MG1655</i> containing both plasmids for a. Western Blots were used to determine expression of <i>ChrB</i>, <i>ChrBopt</i>, and GFP.</a></p>
           </div>
           <div id="collapsiblew9" class="collapse week-content">
@@ Line 1,268: / Line 1,174: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1 - 17/08: Transformations akin to 13/08 were done, but pChrGFP(15/07) and pChrGFP(31/07) were transformed into MG1655+ChrB2 and MG1655+ChrBopt3.</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 17/08: Transformations akin to 13/08 were done, but pSB1C3-pChr-GFP(15/07) and pSB1C3-pChr-GFP(31/07) were transformed into <i>MG1655+pUniprom-ChrB2</i> and <i>MG1655+pUniprom-ChrBopt3</i>.</span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew9-1"></span>
                  <div id="collapsiblew9-1" class="collapse box-content">
@@ Line 1,276: / Line 1,182: @@
                    <p><b>Protocols Used:</b>
-                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformation, </a>
+                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformation </a>
                    </p>
                    <p><b>Results:</b> Successful transformants were found on all plates.</p>
-                  <!-- For images in the results use the following line of code:
+                   <p><b>Next Steps:</b> To determine expression of <i>ChrB</i> and <i>ChrBopt</i> (via 6His-tag) and expression of GFP.</p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#lbpa-figure1-modal" class="journal-protocol"> Figure 1 </a></p>
-                  under data-target refer to a modal specifically made for each image under image modals section-->
-                  <p><b>Next Steps:</b> To determine expression of ChrB and ChrBopt (via His-tag) and expression of GFP.</p>
                  </div>
@@ Line 1,304: / Line 1,203: @@
-                   <p><b>Aim of experiment:</b> To determine whether ChrB, ChrBopt, and GFP are expressed and have first estimates of differences in expression levels.</p>
+                   <p><b>Aim of experiment:</b> To determine whether <i>ChrB</i>, <i>ChrBopt</i>, and GFP are expressed and have first estimates of differences in expression levels.</p>
                    <p><b>Protocols Used:</b>
@@ Line 1,323: / Line 1,222: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 3-6 - 19/08 - 22/08: Repeated western blots with different concentrations to determine whether the expression of ChrB has an effect on the expression of GFP.</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 3-6 - 19/08 - 22/08: Repeated western blots with different concentrations to determine whether the expression of <i>ChrB</i> has an effect on the expression of GFP.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew9-3"></span>
                  <div id="collapsiblew9-3" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> To prepare whether expression of ChrB or ChrB opt has an effect on the expression of GFP.</p>
+                   <p><b>Aim of experiment:</b> To determine whether expression of <i>ChrB</i> or <i>ChrBopt</i> has an effect on the expression of GFP.</p>
                    <p><b>Protocols Used:</b>
@@ Line 1,336: / Line 1,235: @@
                    <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150822_blot" class="journal-protocol"> Figure 28 </a></p>
-                   <p><b>Next Steps:</b> Check if different substances containing chromate have an effect on the expression.</p>
+                   <p><b>Next Steps:</b> Check if different chromate salts have an effect on the respective expression levels.</p>
                  </div>
@@ Line 1,383: / Line 1,282: @@
                    </p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150824_platereader_BBa_K1058008" class="journal-protocol"> Figure 29 </a></p>
+                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150824_platereader_BBa_K1058008" class="journal-protocol"> Figure 29 </a> <i>MG1655 + <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a></i></p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150824_platereader_pchrbgfp+chrb" class="journal-protocol"> Figure 30 </a></p>
+                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150824_platereader_pchrbgfp+chrb" class="journal-protocol"> Figure 30 </a><i>MG1655 + pSB1C3-pChr-GFP + pUniprom-ChrB</i></p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150824_platereader_pchrbgfp+chrbopt" class="journal-protocol"> Figure 31 </a></p>
+                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150824_platereader_pchrbgfp+chrbopt" class="journal-protocol"> Figure 31 </a><i>MG1655 + pSB1C3-pChr-GFP + pUniprom-ChrBopt</i></p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150824_platereader_pchrbgfp" class="journal-protocol"> Figure 32 </a></p>
+                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150824_platereader_pchrbgfp" class="journal-protocol"> Figure 32 </a><i>MG1655 + pSB1C3-pChr-GFP</i></p>
-                   <p><b>Next Steps:</b> Try the system with different expression levels of repressor. Induce overexpression of repressor from T7-promoter in pUniprom using BL21 and IPTG.</p>
+                   <p><b>Next Steps:</b> Try the system with different expression levels of <i>ChrB</i> and <i>ChrBopt</i>. Induce overexpression by exploiting the T7-promoter in pUniprom after transforming the different combinations between pSB1C3-pChr-GFP and pUniprom-ChrB/pUniprom-ChrBopt into <i>BL21(DE3)</i> and adding IPTG.</p>
                  </div>
@@ Line 1,411: / Line 1,310: @@
       <div class="box">
          <div class="labtitle">Week Beginning 31/08</div>
-          <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew11"></span>
            <div class="box-content">
            <p class="journal-summary-heading">Summary</p>
@@ Line 1,439: / Line 1,338: @@
-          <p class="journal-content">Parts from CeBiTec Bielefeld arrived and attempts at cloning were made. The chromate sensing system was transformed into BL21 and some of the plate reader experiments repeated.</a></p>
+          <p class="journal-content">Parts from CeBiTec Bielefeld arrived and attempts at cloning were made. The chromate sensing system was transformed into <i>BL21(DE3)</i> and some of the plate reader experiments repeated.</a></p>
           </div>
           <div id="collapsiblew12" class="collapse week-content">
@@ Line 1,451: / Line 1,350: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 1 - 07/09: Transformation of pSB1K3 into MG1655.</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 1 - 07/09: Transformation of pSB1K3 (Bielefeld) into <i>MG1655</i>.</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew12-1"></span>
                  <div id="collapsiblew12-1" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> The high-copy plasmid arrived at very low concentration. Hence it was transformed in order to produce more of it before digesting it.</p>
+                   <p><b>Aim of experiment:</b> The high-copy plasmid arrived at a concentration of 7 <sup>ng</sup>/<sub>µl</sub>. Hence it was transformed in order to produce more of it before purifying and digesting it.</p>
                    <p><b>Protocols Used:</b>
                      <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformation, </a>
-                     <a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight culture for competent cells </a>
+                     <p><a data-toggle="modal" data-target="#overnightculture-modal" class="journal-protocol"> Overnight culture</a> for new <i>E. coli MG1655</i></p>
                    </p>
                    <p><b>Results:</b> No transformants were found.</p>
-                  <!-- For images in the results use the following line of code:
+                   <p><b>Next Steps:</b> Retry transformation with new competent <i>E. coli MG1655</i>.</p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#lbpa-figure1-modal" class="journal-protocol"> Figure 1 </a></p>
-                  under data-target refer to a modal specifically made for each image under image modals section-->
-                  <p><b>Next Steps:</b> Retry and make more competent MG1655.</p>
                  </div>
@@ Line 1,488: / Line 1,380: @@
-                   <p><b>Aim of experiment:</b> Test transformation of BBa_K1058008 into new MG1655 competent cells and transformation of pSB1K3-plasmid</p>
+                   <p><b>Aim of experiment:</b> Test transformation of BBa_K1058008 into new <i>MG1655</i> competent cells and re-transformation of pSB1K3 for subsequent miniprep.</p>
                    <p><b>Protocols Used:</b>
-                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformation, </a>
+                     <a data-toggle="modal" data-target="#transformation-modal" class="journal-protocol"> Transformation </a>
                    </p>
                    <p><b>Results:</b> Successful test transformation, but no colonies on pSB1K3 plate.</p>
-                  <!-- For images in the results use the following line of code:
+                   <p><b>Next Steps:</b> Use an alternative compatible plasmid for ligation of codon optimised <i>ChrB</i> (Bielefeld).</p>
-                   <p><b>Results:</b><a data-toggle="modal" data-target="#lbpa-figure1-modal" class="journal-protocol"> Figure 1 </a></p>
-                  under data-target refer to a modal specifically made for each image under image modals section-->
-                  <p><b>Next Steps:</b> Use an alternative plasmid.</p>
                  </div>
@@ Line 1,519: / Line 1,404: @@
-                   <p><b>Aim of experiment:</b> Restriction digest of pT7KS with EcoRI and PstI as alternative to pSB1K3. Ligation of digested ChrB (Bielefeld) and transformation of this plasmid. Also transformations of Chromate-sensor plasmids into BL21 (DE3) in the following combinations: <ul>
+                   <p><b>Aim of experiment:</b> Restriction digest of pT7KS with EcoRI and PstI as alternative to pSB1K3. Ligation of digested <i>ChrB</i> (Bielefeld) and transformation of pT7KS-ChrB. Also transformations of Chromate-sensor plasmids into BL21 (DE3) in the following combinations: <ul>
-                                   <li>pChrGFP + ChrB</li>
+                                   <li>pSB1C3-pChr-GFP + pUniprom-ChrB</li>
-                                   <li>pChrGFP + ChrBopt</li>
+                                   <li>pSB1C3-pChr-GFP + pUniprom-ChrBopt</li>
-                                   <li>ChrB</li>
+                                   <li>pUniprom-ChrB</li>
-                                   <li>ChrBopt</li>
+                                   <li>pUniprom-ChrBopt</li>
                                  </ul>
                    </p>
@@ Line 1,533: / Line 1,418: @@
                    </p>
-                   <p><b>Results:</b>Transformations of the plasmid with ChrB (Bielefeld) yielded 12 colonies, but also colonies on the vector control plate. Transformations into BL21 were successful.</p>
+                   <p><b>Results:</b>Transformations of the plasmid with pT7KS-ChrB (Bielefeld) yielded 12 colonies, but also colonies on the vector control plate. Transformations of pSB1C3-pChr-GFP + pUniprom-ChrB, pSB1C3-pChr-GFP + pUniprom-ChrBopt, pUniprom-ChrB, and pUniprom-ChrBopt into <i>BL21(DE3)</i> were successful.</p>
                    <p><b>Results:</b>
-                     <a data-toggle="modal" data-target="#dundee15_chr_150911_chrbbielefeld" class="journal-protocol"> Figure 33 </a>
+                     <p><a data-toggle="modal" data-target="#dundee15_chr_150911_chrbbielefeld" class="journal-protocol"> Figure 33 </a> pT7KS-ChrB (Bielefeld) transformations</p>
-                     <a data-toggle="modal" data-target="#dundee15_chr_150911_bl21" class="journal-protocol"> Figure 34 </a>
+                     <p><a data-toggle="modal" data-target="#dundee15_chr_150911_bl21" class="journal-protocol"> Figure 34 </a> Transformations into <i>BL21(DE3)</i></p>
                    </p>
-                   <p><b>Next Steps:</b> Confirmation of colonies with pT7KS-ChrB by colony PCR. Preparation of recombinant BL21 for plate reader experiments.</p>
+                   <p><b>Next Steps:</b> Confirmation of colonies with pT7KS-ChrB by colony PCR. Preparation of recombinant <i>BL21(DE3)</i> for plate reader experiments.</p>
                  </div>
@@ Line 1,551: / Line 1,436: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 4 - 10/09: Colony PCR of pT7KS.</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 4 - 10/09: Colony PCR of pT7KS-ChrB (Bielefeld).</span> <!--Title of collapsible box-->
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew12-4"></span>
                  <div id="collapsiblew12-4" class="collapse box-content">
-                   <p><b>Aim of experiment:</b> Check if any of the colonies after ligation of ChrB (Bielefeld) into pT7KS were successful despite an unsuccessful control plate.</p>
+                   <p><b>Aim of experiment:</b> Check if any of the colonies after ligation of ChrB (Bielefeld) into pT7KS were successful despite colonies on the control plate.</p>
                    <p><b>Protocols Used:</b>
@@ Line 1,564: / Line 1,449: @@
                    <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_150911_igem050" class="journal-protocol"> Figure 34 </a></p>
-                   <p><b>Next Steps:</b> Prepare recombinant BL21 for plate reader experiments. Attempt new transformation of ChrB (Bielefeld).</p>
+                   <p><b>Next Steps:</b> Prepare transformed <i>BL21(DE3)</i> for plate reader experiments. Attempt new transformation of pT7KS-ChrB (Bielefeld).</p>
                  </div>
@@ Line 1,575: / Line 1,460: @@
         <div class="border-day">
          <div class="box">
-             <span class="box-title">Day 5 - 11/09: Preparation for Plate reader experiment with BL21 (DE3)</span> <!--Title of collapsible box-->
+             <span class="box-title">Day 5 - 11/09: Preparation for Plate reader experiment with <i>BL21(DE3)</i></span>
                <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew12-5"></span>
                  <div id="collapsiblew12-5" class="collapse box-content">
@@ Line 1,607: / Line 1,492: @@
                    <p><b>Protocols Used:</b>
-                     <a data-toggle="modal" data-target="#platereader-modal" class="journal-protocol"> Plate reader </a>
+                     <a data-toggle="modal" data-target="#platereader-modal" class="journal-protocol"> Plate reader experiments</a>
                    </p>
-                   <p><b>Results:</b>add images once analysed.</p>
+                   <p><b>Results:</b><a data-toggle="modal" data-target="#dundee15_chr_platereaderinduced" class="journal-protocol"> Figure 35 </a></p>
                    <p><b>Next Steps:</b>Repeat experiments with added chromate substances.</p>
@@ Line 1,626: / Line 1,511: @@
+<!-- WEEK 13 -->
+  <div class="row">
+    <div class="border-week">
+     <div class="box">
+        <div class="labtitle">Week Beginning 14/09</div>
+         <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew13"></span>
+          <div class="box-content">
+          <p class="journal-summary-heading">Summary</p>
+         <p class="journal-content">During this week the samples from the previous plate reader were retrieved and prepared for blotting against both GFP, and the 6His-Tag of <i>ChrB</i> and <i>ChrBopt</i>.</a></p>
+         </div>
+         <div id="collapsiblew13" class="collapse week-content">
+         <!-- when copying for next week, make sure to change labtitle, span, collapsible in last line. In each separate day change span and following div-->
+      <div class="row">
+       <div class="border-day">
+        <div class="box">
+            <span class="box-title">Day 1 - 14/09: Western blot of plate reader samples <i>MG1655 + pSB1C3-pChr-GFP + pUniprom-ChrB</i>, and  <i>MG1655 + pSB1C3-pChr-GFP + pUniprom-ChrBopt</i>.</span> <!--Title of collapsible box-->
+              <span type="button" class="chevron_toggleable box-btn glyphicon glyphicon-chevron-down" data-toggle="collapse" data-target="#collapsiblew13-1"></span>
+                <div id="collapsiblew13-1" class="collapse box-content">
+                  <p><b>Aim of experiment:</b> To identify whether or not there is any detectable difference in both, the level of GFP, and the level of ChrB or ChrBopt, before and after induction with IPTG.</p>
+                  <p><b>Protocols Used:</b>
+                    <a data-toggle="modal" data-target="#sds-modal" class="journal-protocol"> SDS-PAGE + Western blot </a>
+                  </p>
+                  <p><b>Results:</b> Insert blot image here.</p>
+                  <p><b>Next Steps:</b> Repeat the assays of the previous weeks by subcloning ChrB, ChrBopt, and ChrB (Bielefeld) into vectors with different expression levels and see how it influences the level of GFP expression. Then add different chromate salts and also probe the system with other substances, e.g. other heavy-metal salts.</p>
+                </div>
+        </div>
+        </div>
+      </div>
@@ Line 2,666: / Line 2,591: @@
        <div class="modal-header">
          <button type="button" class="close" data-dismiss="modal" aria-label="close" aria-hidden="true" role="dialog" tabindex="-1"><span aria-hidden="true">&times;</span></button>
-         <div class="modal-title" id="myModalLabel">Figure 2: 1% agarose gel after PCR of <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> with primers specific to the promoter region, pChr, and the repressor region, ChrB. Left: pChr (expected size 169bp); Middle: 1kb+ ladder, Right: ChrB (expected size 939bp).
+         <div class="modal-title" id="myModalLabel">Figure 2: 1% agarose gel after PCR of <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> with primers specific to the promoter region, pChr, and the repressor region, ChrB. Left: pChr (expected size 169bp); Middle: 1kb+ ladder, Right: ChrB (expected size 939bp). Amplification of ChrB was not successful. pChr was further processed by restriction digest with EcoRI and PstI
          </div>
        </div>
@@ Line 2,688: / Line 2,613: @@
        <div class="modal-header">
          <button type="button" class="close" data-dismiss="modal" aria-label="close" aria-hidden="true" role="dialog" tabindex="-1"><span aria-hidden="true">&times;</span></button>
-         <div class="modal-title" id="myModalLabel">Figure 3: 1% agarose gel after PCR of <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> with primers specific to the repressor region, ChrB. Left: 1.5µl DMSO; Middle: 1kb+ ladder, Right: 2.5µl DMSO. Expected size for each: 939bp.
+         <div class="modal-title" id="myModalLabel">Figure 3: 1% agarose gel after PCR of <a href="http://parts.igem.org/Part:BBa_K1058008" target="_blank">BBa_K1058008</a> with primers specific to the repressor region, ChrB. Left: 1.5µl DMSO; Middle: 1kb+ ladder, Right: 2.5µl DMSO. Expected size for each: 939bp. The annealing temperature for the PCR amplification was lowered to 50°C in order to increase the chances of accomodating a primer that is not 100% complementary.
          </div>
        </div>
@@ Line 2,710: / Line 2,635: @@
        <div class="modal-header">
          <button type="button" class="close" data-dismiss="modal" aria-label="close" aria-hidden="true" role="dialog" tabindex="-1"><span aria-hidden="true">&times;</span></button>
-         <div class="modal-title" id="myModalLabel">Figure 4: 1% agarose gel after PCR of GFPmut2 with corresponding primers primers. Left: GFPmut2 (expected size 717bp); Middle: 1kb+ ladder, Right: PCR product of ChrB, using primers to add a BamHI restriction site in front of it and a His-Tag and a HindIII restriction site at the end. (expected size of ChrB: 939bp).
+         <div class="modal-title" id="myModalLabel">Figure 4: 1% agarose gel after PCR of GFPmut2 with corresponding primers primers. Left: GFPmut2 (expected size 717bp); Middle: 1kb+ ladder, Right: PCR product of ChrB, using primers to add a BamHI restriction site in front of it and a His-Tag and a HindIII restriction site at the end. (expected size of ChrB: 939bp). Since amplification of ChrB was unsuccessful, it will be repeated.
          </div>
        </div>
@@ Line 2,825: / Line 2,750: @@
        <div class="modal-body">
         <img src="https://static.igem.org/mediawiki/2015/6/6f/Dundee15_chr_igem035.png" style="width:400px;height:auto">'
+       <!--insert link of uploaded files here-->
+      </div>
+      <div class="modal-footer">
+        <button type="button" class="btn btn-primary" data-dismiss="modal">Close</button>
+      </div>
+    </div>
+  </div>
+</div>
+<div class="modal fade" id="dundee15-chr-igem032-modal" tabindex="-1" role="dialog" aria-labelledby="myModalLabel">
+  <div class="modal-dialog" role="document">
+    <div class="modal-content">
+      <div class="modal-header">
+        <button type="button" class="close" data-dismiss="modal" aria-label="close" aria-hidden="true" role="dialog" tabindex="-1"><span aria-hidden="true">&times;</span></button>
+        <div class="modal-title" id="myModalLabel">Figure 10: 1% agarose gel after PCR of optimised ChrB. (Expected size 939bp).
+        </div>
+      </div>
+      <div class="modal-body">
+       <img src="https://static.igem.org/mediawiki/2015/2/2f/Dundee15_chr_igem032.png" style="width:400px;height:auto">'
         <!--insert link of uploaded files here-->
@@ Line 2,864: / Line 2,811: @@
        <div class="modal-header">
          <button type="button" class="close" data-dismiss="modal" aria-label="close" aria-hidden="true" role="dialog" tabindex="-1"><span aria-hidden="true">&times;</span></button>
-         <div class="modal-title" id="myModalLabel">Figure 12: 1% agarose gel after restriction digest ofChrB-pUniprom, optimised ChrB-pUniprom and pChr-GFPmut2-pSB1C3 for confirmation of size. Ladder: 1kb+. Top: 1-4: pSB1C3-pChr-GFP 1-4; pUniprom-ChrB-opt 1-2. Bottom: pUniprom-ChrB-opt 3-4; pUniprom-ChrB 1-4. Expected sizes: pChr-GFP: 909bp; ChrB and Chrb-opt: 939bp.
+         <div class="modal-title" id="myModalLabel">Figure 12: 1% agarose gel after restriction digest of pUniprom-ChrB, pUniprom-ChrB optimised and pSB1C3-pChr-GFP for confirmation of size. Ladder: 1kb+. Top: 1-4: pSB1C3-pChr-GFP 1-4; pUniprom-ChrB-opt 1-2. Bottom: pUniprom-ChrBopt 3-4; pUniprom-ChrB 1-4. Expected sizes: pChr-GFP: 909bp; ChrB and Chrb-opt: 939bp.
          </div>
        </div>
@@ Line 3,386: / Line 3,333: @@
    </div>
 </div>
+<div class="modal fade" id="dundee15_chr_platereaderinduced" tabindex="-1" role="dialog" aria-labelledby="myModalLabel">
+  <div class="modal-dialog" role="document">
+    <div class="modal-content">
+      <div class="modal-header">
+        <button type="button" class="close" data-dismiss="modal" aria-label="close" aria-hidden="true" role="dialog" tabindex="-1"><span aria-hidden="true">&times;</span></button>
+        <div class="modal-title" id="myModalLabel">Figure 35: Agarose gel after colony PCR.
+        </div>
+      </div>
+      <div class="modal-body">
+       <img src="https://static.igem.org/mediawiki/2015/8/8a/Dundee15_chr_platereaderinduced.png" style="width:400px;height:auto">'
+       <!--insert link of uploaded files here-->
+      </div>
+      <div class="modal-footer">
+        <button type="button" class="btn btn-primary" data-dismiss="modal">Close</button>
+      </div>
+    </div>
+  </div>
+</div>

Difference between revisions of "Team:Dundee/labjournal manu"

Revision as of 18:26, 16 September 2015

BioSpray

Chromium Detector

Fingerprint Aging