Difference between revisions of "Team:Paris Saclay/Notebook/August/4"
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=Tuesday 4th August= | =Tuesday 4th August= | ||
==Lab Work== | ==Lab Work== | ||
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''by Pauline'' | ''by Pauline'' | ||
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* BBa_K1707008 #1 to #6 | * BBa_K1707008 #1 to #6 | ||
* BBa_I13600 #1 and #2 | * BBa_I13600 #1 and #2 | ||
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''by Pauline and Coralie'' | ''by Pauline and Coralie'' | ||
| − | + | * BBa_K1707008 #1 to #6 | |
Mix: | Mix: | ||
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EcoRI + SpeI: | EcoRI + SpeI: | ||
* BBa_K1707004 | * BBa_K1707004 | ||
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Mix: | Mix: | ||
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* 6 µL H2O | * 6 µL H2O | ||
* 10 µL Plasmide | * 10 µL Plasmide | ||
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===Purification agarose gel=== | ===Purification agarose gel=== | ||
''by Coralie and Audrey'' | ''by Coralie and Audrey'' | ||
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* BBa_K1707011 | * BBa_K1707011 | ||
* BBa_K1707004 | * BBa_K1707004 | ||
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We can conclude that I13602 isn't digested | We can conclude that I13602 isn't digested | ||
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===Purification=== | ===Purification=== | ||
''by Pauline'' | ''by Pauline'' | ||
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* BBa_K1707011 | * BBa_K1707011 | ||
* BBa_K1707004 | * BBa_K1707004 | ||
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===Soil experiment=== | ===Soil experiment=== | ||
''by Coralie'' | ''by Coralie'' | ||
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====Water==== | ====Water==== | ||
Sea Water separate in 5mL in 50mL Falcon | Sea Water separate in 5mL in 50mL Falcon | ||
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Water + concentrate cells solution 10 fold dilution | Water + concentrate cells solution 10 fold dilution | ||
Water + concentrate cells solution 100 fold dilution | Water + concentrate cells solution 100 fold dilution | ||
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J0: we take 100µL of each water and put it on plates with specific antibiotic: | J0: we take 100µL of each water and put it on plates with specific antibiotic: | ||
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We choose to only use MCK+Sorbitol now | We choose to only use MCK+Sorbitol now | ||
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===Low Budget Challenge=== | ===Low Budget Challenge=== | ||
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We try another time, with the same protocol than the 03/08/2015 but a fresh culture of bacteria. | We try another time, with the same protocol than the 03/08/2015 but a fresh culture of bacteria. | ||
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'''Member present:''' | '''Member present:''' | ||
Revision as of 07:31, 15 September 2015
Contents
Tuesday 4th August
Lab Work
Plasmid extraction
by Pauline
- BBa_K1707008 #1 to #6
- BBa_I13600 #1 and #2
With the Macherey-Nagel Extraction kit
Digestion Verification
by Pauline and Coralie
- BBa_K1707008 #1 to #6
Mix:
- 2 µL Plasmid
- 0,5 µL XbaI
- 0,5 µL PstI
- 1 µL Buffer FastDigest 10x
- 6 µL H2O
Incubation 2h, 37°C
Electrophoresis
by Pauline
Agarose gel 1% Migration 90V
We can conclude that all BBa_K1707008 clones are OK
We make a glycerol stock of #1 and #2
Electrophoresis Verification
by Pauline and Coralie
Agarose gel, 1%
Migration 100V
We can conclude that:
- BBa_K1707021: #2, #3, #4 and #6 are OK
- BBa_K1707011: #1 to #6 are OK
- BBa_K1707004: #1 to #6 are OK
We make glycerol stock of them.
Digestion
by Coralie
XbaI + PstI:
- BBa_K1707011
- BBa_K1707004 x2
- BBa_E0022
- BBa_E0422
- BBa_K1707012
SpeI + Pst:
- BBa_R0051 x2
- BBa_K1707013
- BBa_K1707019
- BBa_K1707000
XbaI + EcoRI:
- BBa_I13602
EcoRI + SpeI:
- BBa_K1707004
Mix:
- 1 µL Enzyme 1
- 1 µL Enzyme 2
- 2 µL Buffer FastDigest 10x
- 6 µL H2O
- 10 µL Plasmide
Purification agarose gel
by Coralie and Audrey
- BBa_K1707011
- BBa_K1707004
- BBa_E0022
- BBa_E0422
- BBa_K1707012
- BBa_I13602
Agarose gel 1%
Migration 100 V
We can conclude that I13602 isn't digested
Purification
by Pauline
- BBa_K1707011
- BBa_K1707004
- BBa_E0022
- BBa_E0422
- BBa_K1707012
- BBa_R0051
- BBa_K1707013
- BBa_K1707019
- BBa_K1707000
- BBa_K1707004
With Macherey-Nagel Kit
Soil experiment
by Coralie
Water
Sea Water separate in 5mL in 50mL Falcon Fresh water separate in 5mL in 50mL Falcon Contamination with 1mL Water + LB Water + concentrate cells solution Water + concentrate cells solution 10 fold dilution Water + concentrate cells solution 100 fold dilution
J0: we take 100µL of each water and put it on plates with specific antibiotic:
- MacConkey + Spectinomycin
- MacConkey + Tetracyclin
- MacConkey + Chloramphenicol
And only MacConkey for the Water + LB
Incubation ON, 37°C
Soil
Observation of plates (03/08/2015):
Less contamination on plates with MCK+Sorbitol thant MCK+Lactose.
We choose to only use MCK+Sorbitol now
Low Budget Challenge
by Coralie
Observation of plates (03/08/2015): We can saw a little bit of colony. We can suppose that the initial culture wasn't enough in growing phase.
We try another time, with the same protocol than the 03/08/2015 but a fresh culture of bacteria.
Member present:
- Instructors: Claire
- Students: Coralie, Audrey and Pauline
eigem.parissaclay AT gmail DOT com
t@iGEMParisSaclay
aUniversité Paris Sud
a91405 Orsay, France
Copyright © 2015 iGEM Paris Saclay. *This wiki was conceived during long nights of work and a great coffee experience in the beautiful Orsay, France.