Haynes Lab:Notebook/CRISPR Editing/2015/05/17

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05/17/2015

Added 1ul enhancer and 1ul surveyor enzyme to the 11 tubes I annealed yesterday. 2ul of water to the surveyor negative tubes. incubated at 42°C for 60 mins, added 2.5ul stop solution, ran on bioanalyzer

ligation plates from yesterday

DNA Cells # colonies
lig mine 24
bb mine 14
- mine 17
lig KAH 7
bb KAH 15
- KAH 0

Purified the digests with Qiaquick PCR purification kit. Eluted in 30ul. Will retry the ligation tomorrow. Will order more of the zymo competent cell mix

Sample Read# 260 280 260/280 ng/µL
insert -0.001 0 ????? -0.622
Backbone 0.007 0.004 1.775 7.424

Should recut more backbone, do like 2-3 reactions for a total of 2-3ug, column purify. Then cut the insert and dephos it. Should also make sure the EcoRI works in cutsmart buffer.