Haynes Lab:Notebook/CRISPR Editing/2014/09/21

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09/21/2014

Ran the PCR for gRNA 20 samples from yesterday on a gel. Ran 3μL of each sample and 2μL and 1μL of ladder. Only the first three worked, even though the other two worked the other day.

Annealing all of each of gRNA 20, plus the PCRs from the other day that worked. Will just go through the protocol to see if I can even do Surveyor with the concentrations I'm getting.



Going to try more PCR. Will do gRNA 21 samples and all the Luc14 samples. If I can get all the Luc14 samples through the entire protocol, I can know if any of the gRNAs aren't cutting at all and won't have to keep trying the chromatin-induced samples.



transferred 40ul of the first three annealing reactions and 35ul of the last two to new tubes. added 1ul enhancer and 2ul of surveyor nuclease. incubate at 42deg C for 1 hour.
added 5ul of water to whatever is left in the PCR tubes from annealing. will run those as a no-nuclease control.


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