Haynes Lab:Notebook/CRISPR Editing/2015/01/31

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01/30/2015

miniprepped colonies from topo cloning plate for g034

Sample Read# 260 280 260/280 ng/µL
Topo clone 1 0.02 0.011 1.827 19.983
Topo clone 2 0.057 0.03 1.882 57.143
Topo clone 3 0.582 0.302 1.924 581.887
Topo clone 4 0.018 0.008 2.333 17.68
Topo clone 5 0.046 0.025 1.851 46.218
Topo clone 6 0.04 0.021 1.896 40.046


Ran gradient PCR reactions from yesterday on gel. Setting AT of 66 as 100%, estimated the drop in efficiency. Def not quantitative. There was an obvious off target band around 800bp that faded and disappeared at higher annealing temperatures.

Annealing Temps Efficiency off target band ~800bp
66 100 yes
67 100 yes
68 100 yes
69 80 faint to gone
70 80 gone
72 60 gone


Split Cameron's cells into two T-25 flasks.