Haynes Lab:Notebook/CRISPR Editing/2014/09/21: Difference between revisions
Rene M Davis (talk | contribs) |
Rene M Davis (talk | contribs) |
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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. | 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. | ||
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{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Set 1''' | |||
| align="center" style="background:#f0f0f0;"|'''Set 2''' | |||
|- | |||
| gRNA 21 + dox 1||gRNA 20 Luc 14 | |||
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| gRNA 21 + dox 2||gRNA 21 Luc 14 | |||
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| gRNA 21 - dox 1||gRNA 22 Luc 14 | |||
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| gRNA 21 - dox 2||gRNA 27 Luc 14 | |||
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| gRNA 21 Luc 14||gRNA 29 Luc 14 | |||
|} | |||
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Master mix 1 | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''''' | |||
| align="center" style="background:#f0f0f0;"|'''1rxn''' | |||
| align="center" style="background:#f0f0f0;"|'''10.5 rxns''' | |||
|- | |||
| H20||23.5||246.75 | |||
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| 5x buffer||10||105 | |||
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| dNTPs||1||10.5 | |||
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| FP||2.5||26.25 | |||
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| RP||2.5||26.25 | |||
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| DMSO||1.5||15.75 | |||
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| ||41|| | |||
|} | |||
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add 41ul of MM 1 to each tube<br> | |||
add 4ul of each template to each tube (400ng each) | |||
Master mix 2 | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''''' | |||
| align="center" style="background:#f0f0f0;"|'''1rxn''' | |||
| align="center" style="background:#f0f0f0;"|'''10.5 rxns''' | |||
|- | |||
| H20||4.5||47.25 | |||
|- | |||
| Phusion||0.5||5.25 | |||
|} | |||
add 5 ul of MM 2 to each tube | |||
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Revision as of 14:59, 21 September 2014
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09/21/2014Ran 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.
add 5 ul of MM 2 to each tube
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