Difference between revisions of "Team:Paris Saclay/Notebook/July/11"

(→‎Electrocompetents and transformation by electroporation of BL21)
(→‎Monday 11th July)
Line 11: Line 11:
 
*BL21|K1372001+pcl_TAG
 
*BL21|K1372001+pcl_TAG
 
*BL21|K1372001+pcl_Tq
 
*BL21|K1372001+pcl_Tq
 +
 +
 +
===Bringing DNA closer===
 +
====DH5α transformation with ST1, NM, SP and TD====
 +
''By Laetitia and Mathilde''
 +
 +
DS-NMcas, DS-SPcasN-, DS-ST1cas-N and DS-TDcasN- plasmids were resuspended with 5μL of sterilized water.
 +
5 tubes were prepared :
 +
*one control tube with only 50μL of DH5α heat-shock competent cells
 +
*four tubes with 50μL of DH5α heat-shock competent cells + 1μL of plasmid
 +
 +
The regular heat-shock competent cells [[Team:Paris_Saclay/Experiments#HeatShockCompetent|transformation protocol]] was used.
 +
 +
Petri dishes preparation :
 +
*200mL of LB Agar
 +
*130μL (50μL/mL) of spectinomycin   
 +
Transformed cells were plated in duplicata (10μL and 50μL) on 8 plates. Control cells were plated on another plate.

Revision as of 16:40, 11 July 2016

Monday 11th July

Lab work

Biobrick characterization

Electrocompetents and transformation by electroporation of BL21

By Charlène

BL21 cells were grown up to OD600nm=7.14. They were then made competent and transformed with K1372001 extracted from clone 2 and pcl_TAA, pcl_TAG or pcl_Tq to create the following strains:

  • BL21|K1372001+pcl_TAA
  • BL21|K1372001+pcl_TAG
  • BL21|K1372001+pcl_Tq


Bringing DNA closer

DH5α transformation with ST1, NM, SP and TD

By Laetitia and Mathilde

DS-NMcas, DS-SPcasN-, DS-ST1cas-N and DS-TDcasN- plasmids were resuspended with 5μL of sterilized water. 5 tubes were prepared :

  • one control tube with only 50μL of DH5α heat-shock competent cells
  • four tubes with 50μL of DH5α heat-shock competent cells + 1μL of plasmid

The regular heat-shock competent cells transformation protocol was used.

Petri dishes preparation :

  • 200mL of LB Agar
  • 130μL (50μL/mL) of spectinomycin

Transformed cells were plated in duplicata (10μL and 50μL) on 8 plates. Control cells were plated on another plate.