Team:Paris/August 27
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(Difference between revisions)
(→Measurement of the concentration of D187 purified) |
(→Construction of pFlhB - mRFP Tripart (LVA+)) |
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[[Team:Paris/Notebook/Protocols#Concentration_of_the_Miniprep | Protocol (it's same that for Miniprep)]] | [[Team:Paris/Notebook/Protocols#Concentration_of_the_Miniprep | Protocol (it's same that for Miniprep)]] | ||
- | => the experiments have failed, so we need to repeat the step of digestion. | + | => the experiments of Gel Extraction have failed, so we need to repeat the step of digestion. |
+ | |||
+ | |||
+ | |||
+ | ===Digestion=== | ||
+ | [[Team:Paris/Notebook/Protocols#Digestion|Protocol Digestion]] | ||
+ | |||
+ | {| border="1" style="text-align: center" | ||
+ | |'''Name''' | ||
+ | |'''Template DNA''' | ||
+ | |'''Description''' | ||
+ | |'''Vol MP (µl)''' | ||
+ | |'''Vol H2O (µl)''' | ||
+ | |'''Enzymes''' | ||
+ | |- | ||
+ | |D187 | ||
+ | |MP168.1 | ||
+ | | RBS - mRFP - term (FV) | ||
+ | |9.00 | ||
+ | |15.7 | ||
+ | |EcoRI and XbaI | ||
+ | |} | ||
+ | |||
+ | |||
+ | ==='''Gel Verification'''=== | ||
+ | [[Team:Paris/Notebook/Protocols#Migration after extraction |Protocol]] | ||
+ | [[Image:KR000.JPG |thumb |Gel Verification of D187 digestion]] | ||
+ | |||
+ | {|border="1" style="text-align: center" | ||
+ | |'''Well''' | ||
+ | |1 | ||
+ | |2 | ||
+ | |3 | ||
+ | |4 | ||
+ | |5 | ||
+ | |6 | ||
+ | |- | ||
+ | |'''Sample''' | ||
+ | |100 pb ladder | ||
+ | |MP168.1 | ||
+ | |no sample | ||
+ | |D187 | ||
+ | |colspan="2"|no sample | ||
+ | |- | ||
+ | |'''Expected size (pb)''' | ||
+ | | | ||
+ | | | ||
+ | | | ||
+ | | | ||
+ | | colspan="2"| | ||
+ | |- | ||
+ | |'''Measured size (pb)''' | ||
+ | | | ||
+ | |style="background: #cbff7B"| | ||
+ | | | ||
+ | |style="background: #cbff7B"| | ||
+ | |colspan="2"| | ||
+ | |} | ||
=Cloning of EnvZ* in pSB1A2= | =Cloning of EnvZ* in pSB1A2= |
Revision as of 18:33, 27 August 2008
Construction of pFlhB - mRFP Tripart (LVA+)Aim : Construction of "pFlhB-RBS-mRFP-dbl ter" (pFlhB-I732078) We can only do the construction with mRFP Tripart (LVA+) because the stable strain with the Biobricks I732011 (mRFP Tripart LVA-) don't to growth.
DigestionMeasurement of the concentration of D187 purifiedProtocol (it's same that for Miniprep) => the experiments of Gel Extraction have failed, so we need to repeat the step of digestion.
Digestion
Gel Verification
Cloning of EnvZ* in pSB1A2Transformation results
PCR screening
Electrophoresis
PCR
elongation time: 2 min 30
Cloning of OmpR*DigestionDetermination of the concentration of DNAWe used the biophotometer
Name of the digestions
Protocol of digestion
Cleaning of the digestion productsLigationDetermination of the concentration of DNAWe used the biophotometer
Protocol of ligation L171
Checking mutagenesis FliAFor this, i digested mutated FliA and non-mutated FliA with EcoRI and PstI and put in migration the digestion products running on gels. |