Haynes Lab:Notebook/CRISPR Editing/2016/05/27: Difference between revisions
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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span> | |style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;"> Project name</span> | ||
|style="background-color: #F2F2F2" align="center"| | |style="background-color: #F2F2F2" align="center"|[[File:Report.png|frameless|link={{#sub:{{FULLPAGENAME}}|0|-11}}]][[{{#sub:{{FULLPAGENAME}}|0|-11}}|Main project page]]<br />{{#if:{{#lnpreventry:{{FULLPAGENAME}}}}|[[File:Resultset_previous.png|frameless|link={{#lnpreventry:{{FULLPAGENAME}}}}]][[{{#lnpreventry:{{FULLPAGENAME}}}}{{!}}Previous entry]] }}{{#if:{{#lnnextentry:{{FULLPAGENAME}}}}|[[{{#lnnextentry:{{FULLPAGENAME}}}}{{!}}Next entry]][[File:Resultset_next.png|frameless|link={{#lnnextentry:{{FULLPAGENAME}}}}]]}} | ||
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<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### --> | <!-- ##### DO NOT edit above this line unless you know what you are doing. ##### --> | ||
== | ==05/27/2016== | ||
Ran g032, g044, g046 P197/198 PCR from yesterday on gel. G032 looked great, g044 and 46 were very faint. Ran nested PCR of all of them. G032 diluted 1:1000, dox with my phusion, puro with braflab phusion, 44 and 46 diluted 3:1000 with braflab phusion. | |||
<br><br> | |||
Extracted gDNA from KAH228/g034 cells: | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Cell type''' | |||
| align="center" style="background:#f0f0f0;"|'''drug''' | |||
| align="center" style="background:#f0f0f0;"|'''re-opener''' | |||
| align="center" style="background:#f0f0f0;"|'''gRNA''' | |||
| align="center" style="background:#f0f0f0;"|'''rep''' | |||
| align="center" style="background:#f0f0f0;"|'''260''' | |||
| align="center" style="background:#f0f0f0;"|'''280''' | |||
| align="center" style="background:#f0f0f0;"|'''260/280''' | |||
| align="center" style="background:#f0f0f0;"|'''ng/µL''' | |||
|- | |||
| Luc14||||KAH228||g034||1||0.302||0.151||2.003||301.829 | |||
|- | |||
| Luc14||||KAH228||g034||2||0.652||0.326||1.999||651.792 | |||
|- | |||
| Luc14||||KAH228||g034||3||0.449||0.223||2.009||448.873 | |||
|- | |||
| Gal4||dox||KAH228||g034||1||0.335||0.167||2||334.767 | |||
|- | |||
| Gal4||dox||KAH228||g034||2||0.333||0.166||2.006||333.159 | |||
|- | |||
| Gal4||dox||KAH228||g034||3||0.305||0.153||1.987||304.635 | |||
|} | |||
<br><br> | |||
Nested PCR of g032, g044, g046<br> | |||
:Gal4 +puro g032 had faint bands, no band in neg ctrl, decided to purify | |||
:Gal4 +dox g034 had no bands, my phusion is bad | |||
:Luc14 g044 faint bands, no band in neg ctrl, purified | |||
:Luc14 g046 super bright bands, no band in neg ctrl | |||
<br> | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Cell type''' | |||
| align="center" style="background:#f0f0f0;"|'''drug''' | |||
| align="center" style="background:#f0f0f0;"|'''gRNA''' | |||
| align="center" style="background:#f0f0f0;"|'''rep''' | |||
| align="center" style="background:#f0f0f0;"|'''260''' | |||
| align="center" style="background:#f0f0f0;"|'''280''' | |||
| align="center" style="background:#f0f0f0;"|'''260/280''' | |||
| align="center" style="background:#f0f0f0;"|'''ng/µL''' | |||
| align="center" style="background:#f0f0f0;"|'''ul for 400ng''' | |||
| align="center" style="background:#f0f0f0;"|'''ul ann buff''' | |||
| align="center" style="background:#f0f0f0;"|'''ul H2O''' | |||
|- | |||
| Gal4||puro||g032||1||0.016||0.009||1.86||16.007||24.99||3.50||6.51 | |||
|- | |||
| Gal4||puro||g032||2||0.008||0.004||2.116||8.125||49.23|||| | |||
|- | |||
| Gal4||puro||g032||3||0.013||0.007||1.812||12.543||31.89||3.50||-0.39 | |||
|- | |||
| Luc14||||g044||1||0.015||0.008||1.763||14.617||27.37|||| | |||
|- | |||
| Luc14||||g044||2||0.009||0.006||1.552||9.059||44.15|||| | |||
|- | |||
| Luc14||||g044||3||0.056||0.03||1.864||56.272||7.11||1.50||6.39 | |||
|- | |||
| Luc14||||g046||1||0.11||0.058||1.877||109.619||3.65||1.50||9.85 | |||
|- | |||
| Luc14||||g046||2||0.094||0.05||1.877||93.996||4.26||1.50||9.24 | |||
|- | |||
| Luc14||||g046||3||0.102||0.055||1.843||101.658||3.93||1.50||9.57 | |||
|} | |||
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Latest revision as of 01:48, 27 September 2017
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05/27/2016Ran g032, g044, g046 P197/198 PCR from yesterday on gel. G032 looked great, g044 and 46 were very faint. Ran nested PCR of all of them. G032 diluted 1:1000, dox with my phusion, puro with braflab phusion, 44 and 46 diluted 3:1000 with braflab phusion.
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