From 2009.igem.org
(Difference between revisions)
|
|
| Line 26: |
Line 26: |
| | | | |
| | ==Wet Lab== | | ==Wet Lab== |
| - | ===Colony PCR===
| |
| - | On the same double transformants -> 2 min extension.
| |
| - |
| |
| - | ===Gel===
| |
| - | Here the 2nd strains seems to have a double transformant. But we let the gel overnight so there is no BrEth any more, we had to take a time of exposure of 38.2 s to see something. We will do a nicer gel this afternoon.
| |
| - |
| |
| - | ===Preparation of the time-course experiment===
| |
| - | The idea is to look at the evolution of fluorescence over the time, for different time of exposition on the light.
| |
| - |
| |
| - | 7 ml of LB + 1 ml of DH5a RO2.4 + BB1 #3 overnight culture.
| |
| - |
| |
| - | 5 conditions :
| |
| - |
| |
| - | - + light + IPTG - Trp
| |
| - |
| |
| - | - + light - IPTG - Trp
| |
| - |
| |
| - | - - light + IPTG - Trp
| |
| - |
| |
| - | - - light - IPTG + Trp
| |
| - |
| |
| - | - - light - IPTG + Trp
| |
| - |
| |
| - | - - light - IPTG - Trp
| |
| - |
| |
| - | IPTG at 1 mM (100 mM sol. diluted 1/100). Trp diluted 1/25. Incubated in conditions at 37°C. Measures will be taken with plate reader every 30 minutes starting at 1h.
| |
| - |
| |
| - | ===Miniprep===
| |
| - | We did a miniprep of new strain double transformants to confirm gel results by digestion assay :
| |
| - |
| |
| - | - RO1.1 + BB1 JRG 1046 (n°1,3,7)
| |
| - |
| |
| - | - RO2.4 + BB1 JRG 1046 (n°1,3,6)
| |
| - |
| |
| - | + miniprep of DH5 a RO2.4 + BB1 #3 (strain that works) to have the 2 plasmids for subsequent transformations.
| |
| - |
| |
| - | ===Digestion assay===
| |
| - | For each clones, we use digestion by P and S (since LovTAP contains 2 P sites and Trp op contains 2 S sites). For reaction, incubate at 37°C (INCUB37). Digestion of about 1h30.
| |
| - |
| |
| - | ===Gel===
| |
| - | Once again to see the double transformants + of digestion assay.
| |
| - |
| |
| - |
| |
| - | Result ?? TO BE COMPLETED
| |
| | | | |
| | ==People in the lab== | | ==People in the lab== |
Revision as of 07:33, 8 October 2009
8 October 2009
Wet Lab
People in the lab
Heidi, Gab, Tu
"
