Haynes Lab:Notebook/HPK-CFP insertion into Gal4EED/Luc using CRISPR/2015/08/05

Sequence verification around tk-luc (cont'd)

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Overview Trying to repeat the PCR for the first reaction using P197 & P215 primers (René's). PCR Reaction 1 (anneal temp. 54°C): Reagent Volume (µL) 2x GoTaq MM 25 MB grade H2O 24 P197 (RD) 10 µM 0.25 P215 (RD) 10 µM 0.25 template 0.5 Step Temp. (°C) Duration (sec) Initial Denature 95 120 Main cycle (x35): denature 95 30 anneal 46-->54 30 extension 72 60 final extension 72 300 soak 4 indef. Gel Electrophoresis Results Faint but visible bands at all temperature ranges (46-->54). Must have not put in one of the primers during the previous run. Will perform PCR clean-up and submit samples for sequencing in a couple days.