Difference between revisions of "Team:Paris Saclay/Notebook/August/5"
(→PCR Clean-up with the NucleoSpin kit) |
(→Lab work) |
||
| (8 intermediate revisions by 5 users not shown) | |||
| Line 1: | Line 1: | ||
{{Team:Paris_Saclay/notebook_header}} | {{Team:Paris_Saclay/notebook_header}} | ||
| − | = Friday 5<sup> | + | = Friday 5<sup>th</sup> August= |
| − | + | ||
| − | === | + | ===Bringing DNA Closer=== |
==== DNA Extraction of DS-TDcasN- and DS-SPcasN- ==== | ==== DNA Extraction of DS-TDcasN- and DS-SPcasN- ==== | ||
''By Laetitia'' | ''By Laetitia'' | ||
| + | The extraction was performed using the ChargeSwitc-Pro MiniPrep kit from Invitrogen. We followed this [[Team:Paris_Saclay/Experiments#DNA_extraction_using_the_Invitrogen_ChargeSwitch.C2.AE-Pro_Plasmid_Miniprep_Kit|protocol]]. | ||
| − | + | The initial culture was at 20 mL so we increased (*4) the volumes until the precipitation. | |
| − | + | ||
| − | The initial culture was at | + | |
Hence we used: | Hence we used: | ||
| − | + | * 1 ml of resuspension buffer | |
| − | + | * 1 ml lysis buffer | |
| − | + | * 1 ml precipitation buffer | |
| − | + | ||
| − | + | ||
The colum was used several times in order to recover the maximum of DNA | The colum was used several times in order to recover the maximum of DNA | ||
| Line 24: | Line 21: | ||
''By Mathilde'' | ''By Mathilde'' | ||
| − | The extraction was performed on clone 11 for NM and clone 9 for 1.1 using the kit. We followed this [[Team:Paris_Saclay/Experiments#DNA_extraction_using_the_Invitrogen_ChargeSwitch.C2.AE-Pro_Plasmid_Miniprep_Kit|protocol]]. | + | The extraction was performed on clone 11 for NM and clone 9 for 1.1 using the ChargeSwitc-Pro MiniPrep kit from Invitrogen. We followed this [[Team:Paris_Saclay/Experiments#DNA_extraction_using_the_Invitrogen_ChargeSwitch.C2.AE-Pro_Plasmid_Miniprep_Kit|protocol]]. |
| + | ===Visualization=== | ||
==== Q5 PCR on pPS16_003, pPS16_004, pPS16_006 and pPS16_007 ==== | ==== Q5 PCR on pPS16_003, pPS16_004, pPS16_006 and pPS16_007 ==== | ||
''By Caroline'' | ''By Caroline'' | ||
| − | The PCR was carried out following the usual [[Team:Paris_Saclay/Experiments#Q5PCR|protocol]] adapted to 50µL and with a TM at 70°C. The specific [[Team:Paris_Saclay/Experiments#primers|primers]] for each parts were | + | The PCR was carried out following the usual [[Team:Paris_Saclay/Experiments#Q5PCR|protocol]] adapted to 50µL and with a TM at 70°C. The specific [[Team:Paris_Saclay/Experiments#primers|primers]] for each parts were used. The products were put for migration on a 0.8%agarose gel with BET. Only, PCR form pPS16_003 and pPS16_004 showed a positive result. |
| + | |||
| + | |||
| + | [[File:T--Paris Saclay--160805 gel pcr.JPG|400px|thumb|right|Result of the PCR]] | ||
==== PCR Clean-up with the NucleoSpin kit ==== | ==== PCR Clean-up with the NucleoSpin kit ==== | ||
''By Caroline'' | ''By Caroline'' | ||
| − | The purification was carried out on | + | The purification was carried out on PCR products 2.1 and 2.2 following the usual [[Team:Paris_Saclay/Experiments#Purification|protocol]]. |
| − | + | ||
File:T--Paris_Saclay--160722_Visualisation_PCRpPS16_002_7.JPG | File:T--Paris_Saclay--160722_Visualisation_PCRpPS16_002_7.JPG | ||
{{Team:Paris_Saclay/notebook_footer}} | {{Team:Paris_Saclay/notebook_footer}} | ||
Latest revision as of 16:21, 9 October 2016
