iGEM Kent 2015
Notebook
| June |
| M | T | W | T | F | S | S |
| 1 | 2 | 3 | 4 | 5 | 6 | 7 |
| 8 | 9 | 10 | 11 | 12 | 13 | 14 |
| 15 | 16 | 17 | 18 | 19 | 20 | 21 |
| 22 | 23 | 24 | 25 | 26 | 27 | 28 |
| 29 | 30 | | | | | |
| July |
| M | T | W | T | F | S | S |
| | | 1 | 2 | 3 | 4 | 5 |
| 6 | 7 | 8 | 9 | 10 | 11 | 12 |
| 13 | 14 | 15 | 16 | 17 | 18 | 19 |
| 20 | 21 | 22 | 23 | 24 | 25 | 26 |
| 27 | 28 | 29 | 30 | 31 | | |
| August |
| M | T | W | T | F | S | S |
| | | | | | 1 | 2 |
| 3 | 4 | 5 | 6 | 7 | 8 | 9 |
| 10 | 11 | 12 | 13 | 14 | 15 | 16 |
| 17 | 18 | 19 | 20 | 21 | 22 | 23 |
| 24 | 25 | 26 | 27 | 28 | 29 | 30 |
| 31 | | | | | | |
| September |
| M | T | W | T | F | S | S |
| | 1 | 2 | 3 | 4 | 5 | 6 |
| 7 | 8 | 9 | 10 | 11 | 12 | 13 |
| 14 | 15 | 16 | 17 | 18 | 19 | 20 |
| 21 | 22 | 23 | 24 | 25 | 26 | 27 |
| 28 | 29 | 30 | | | | |
Day 1 Wet lab 22/06/15
Made LB plates
Made LB broth
TIPS autoclaved
Day 2 Wet lab 23/06/15
Set up Top10 overnight, VS45 overnight, US348 overnight
Meeting with advisors to discuss progress
Day 3 Wet lab 24/06/15
Filter sterilise the buffer
250ml no antibiotic broth, put top10 cells in
Incubate until optical density is 0.6
Miniprep PSB1C3 plasmid
Day 4 Wet lab 25/06/15
Transformation
Prepare SBC media
Digest of PSB1c3 from MS348
Day 5 Wet lab 26/06/15
Calculated competent cell efficiency
Agarose gel formation
Gel electrophoresis
Day 6 Wet lab 29/06/15
Transforming linear PSB1C3
Day 7 Wet lab 30/06/15
Miniprep of PSB1A3 plasmid
Gel electrophoresis of a large quantity of PSB1C3
Meeting with advisors to discuss progress
Day 8 Wet lab 01/07/15
Gel electrophoresis of the purified DNA extracted
Transformation of pSB1A3 circular plasmid to VS45 cells (competent)
Digest of all pSB1C3 plasmids
Day 9 Wet lab 02/07/15
Competent cells transformation efficiency
Mini-prep of 1MS340 to get pSB1C3 plasmid
Gel extraction of big digest
Quantification of the digest. Added sample buffer to each digest
Day 10 Wet lab 03/07/15
Transformation efficiency
Day 11 Wet lab 06/07/15
TOP10 cells containing pSB1A3 with limonene synthase were cultured overnight on AMP plates
Colonies of VS45 containing pSB1A3 were patched onto chloramphenicol plates
Overnight digest of miniprepped pSBIC3 using ECORI and PSTl
Day 12 Wet lab 07/07/15
An agarose gel of the overnight digest was ran, but no bands were visible
Set up overnight cultures of pSBIC3 and pSBIA3 to be miniprepped the next day
Set up an overnight digest of pSBIC3
Day 13 Wet lab 08/07/15
Agarose gel of overnight digest was ran
miniprep of pSBIA3 and pSBIC3
Day 14 Wet lab 09/07/15
Agarose gel of digested pSBIA3 and pSBIC3 (no bands visible)
Overnight cultures of MS349 containing pSBIA3 LIMS and MS340 containing pSBIC3 were set up, ready to be miniprepped
Day 15 Wet lab 10/07/15
Miniprep of plasmids pSBIA3 and pSBIC3
Day 16 Wet lab 13/07/15
Overnight cultures of MS349 containing pSBIA3 LIMS and MS340 containing pSBIC3 were set up, ready to be miniprepped
Overnight digest of miniprepped plasmids pSBIA3 and pSBIC3 using the enzymes ECORI and PSTI
Day 17 Wet lab 14/07/15
Agarose gel of digested plasmids pSBIA3 and pSBIC3 - it finally worked!!!
Miniprep of overnight cultures
Gel extraction of both plasmids
Analytical gel using Hyperladder I to quantify the plasmid
