Haynes Lab:Notebook/CRISPR Editing/2015/03/03

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03/03/2015

Checked concentrations on minipreps from last year of Lonza pMax GFP from Krishna. very low so JW transformed the original along with pMax GFP 6.2 and KAH60.

Sample Read# 260 280 260/280 ng/µL
pMax GFP 0.026 0.015 1.736 26.294
pMax GFP 0.023 0.013 1.766 23.175
pMax GFP 0.031 0.019 1.639 30.851



Ran PCR products from P151/158 on gel. Only single product for each band.

Sequencing:
Need to purify. Will purify 25ul and elute into 35 ul.
Purification results:

Cell type treatment rep 260 280 260/280 ng/µL
Luc14 Untreated 1 0.035 0.017 2.006 35.007
Luc14 g046 1 0.032 0.016 2.02 31.691
Luc14 Untreated 2 0.045 0.022 2.034 44.797
Luc14 Untreated 3 0.046 0.024 1.938 46.277
Gal4-EED +dox Untreated 1 0.04 0.019 2.099 40.212
Gal4-EED +dox Untreated 2 0.043 0.023 1.885 42.773
Gal4-EED +dox Untreated 3 0.044 0.023 1.93 43.675
Luc14 g046 2 0.03 0.015 2.086 30.259
Luc14 g046 3 0.031 0.015 2.113 31.368
Gal4-EED +dox g046 1 0.035 0.017 2.042 35.122
Gal4-EED +dox g046 2 0.039 0.019 2.05 38.501
Gal4-EED +dox g046 3 0.041 0.022 1.897 41.336
Luc14 Untreated 1 0.032 0.016 1.949 31.937
Luc14 g046 1 0.03 0.015 1.94 29.645


Sent for sequencing at biodesign with P176

Label Cell type treatment rep PCR primer
1 Luc14 Untreated 1 P176
2 Luc14 Untreated 2 P176
3 Luc14 Untreated 3 P176
4 Gal4-EED+dox Untreated 1 P176
5 Gal4-EED+dox Untreated 2 P176
6 Gal4-EED+dox Untreated 3 P176
7 Luc14 g034 1 P176
8 Luc14 g034 2 P176
9 Luc14 g034 3 P176
10 Gal4-EED+dox g034 1 P176
11 Gal4-EED+dox g034 2 P176
12 Gal4-EED+dox g034 3 P176
13 Luc14 g046 1 P176
14 Luc14 g046 2 P176
15 Luc14 g046 3 P176
16 Gal4-EED+dox g046 1 P176
17 Gal4-EED+dox g046 2 P176
18 Gal4-EED+dox g046 3 P176



Ran PCR with P151/158 on g023, g045, g048 PCR with P197/198 diluted 1:1000 from yesterday. Ran on gel.