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

Plasmid Verification (multiple)

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Overview All six liquid cultures grew as expected. Harvested cells and performed a plasmid miniprep for each. The BioBrick cloning constructs ( from KAH57 & KAH104) I'm naming DBN006. DNA Concentration Sample ID Conc. (ng/µL) St Dev (ng/µL) 260/280 DBN006 1 383.4 3.20 1.94 DBN006 2 671.2 11.91 1.93 DBN006 3 298.6 7.32 1.94 KAH57 259.3 3.25 1.93 KAH104 293.7 44.45 1.91 pMaxGFP 549.5 7.52 1.92 Will verify via restriction digest tomorrow. Restriction Digest (05/06) Part Conc. (ng/µL) Volume for 600ng (µL) Volume water (µL) DBN006 1 383.4 1.6 23.4 DBN006 2 671.2 0.9 24.1 DBN006 3 298.6 2.0 23.0 KAH57 259.3 2.3 22.7 KAH104 293.7 2.0 23.0 Reagent Volume (µL) DNA + water 25 10x FD Buffer (Green) 3 EcoRI 1 SpeI 1 Total 30 Incubate at 37°C for 10 minutes, then run total volume on the gel. Digest Results Colonies 1 and 3 contain inserts of the expected size.