Revision as of 19:07, 29 June 2009

Week from June 29th, to June 30th, 2009

We read that using primers VR/VF2 to PCR B0010 will result in excess bands, as documented by Samantha Burke in http://partsregistry.org/Problems_with_PCR_using_VR/VF2 . Maybe our extra bands were due to unwanted annealing between VR primer and B0015 which contains B0010.

In the future we will perform the screening of the ligations either through digestion or colony PCR taking carefully into account of their expected length and their potential non-specific primer binding sites.

We picked 4 colonies from A8 plate and 10 colonies from A9 plate (both stored at +4°C) and infected 1 ml of LB + Amp. We incubated the cultures at 37°C, 220 rpm for 5 and 1/2 hours.

We re-filled the remaining 250 ul of bacterial culture with 5 ml of LB + Amp and incubated the cultures overnight at 37°C, 220 rpm. Tomorrow we will repeat the screening on these plates through miniprep/digestion!

Digestion results for A8 and A9:

@@ Line 30: / Line 30: @@
 *We picked 4 colonies from A8 plate and 10 colonies from A9 plate (both stored at +4°C) and infected 1 ml of LB + Amp. We incubated the cultures at 37°C, 220 rpm for 5 and 1/2 hours.
-*Glycerol stocks for the grown cultures.
+*The 9th picked colony of A9 didn't show any growth.
+*Glycerol stocks for the 13 grown cultures.
 *We re-filled the remaining 250 ul of bacterial culture with 5 ml of LB + Amp and incubated the cultures overnight at 37°C, 220 rpm. Tomorrow we will repeat the screening on these plates through miniprep/digestion!