Haynes Lab:Notebook/HPK-CFP insertion into Gal4EED/Luc using CRISPR/2016/01/08
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Running colony PCR on the HA2_v0120 ligation from yesterday and planning future steps.
Never seen this before... going to pick 8 colonies from the treatment and perform colony PCR using the v0120 primers.
Prepare the following master mix in the first tube of an 8-tube PCR strip:
Mix all ingredients thoroughly then dispense 20 µL into each tube in the strip. Pick 6 colonies from the sample plate, and 2 colonies from the negative plate, using a fresh P20 tip for each one, and place in the reaction mix. Let sit for 10 minutes, then streak an LB-Amp plate split into 8 sections.
Colony PCR program:
Expected product size is 358 bp.
Looks like all 6 colonies picked from the ligation plate worked. Neither of the 2 negative control colonies show amplification in the expected size range. Next step: start liquid cultures from streak plate, plasmid miniprep, move on to next stage of assembly.