Haynes Lab:Notebook/CRISPR Editing/2014/09/23: Difference between revisions
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Rene M Davis (talk | contribs) (Autocreate 2014/09/23 Entry for Haynes_Lab:Notebook/CRISPR_Editing) |
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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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== | ==09/23/2014== | ||
Running 2ul of each PCR rxn on a gel, with 3ul of water and 1ul of 6x loading dye. | |||
<br> | |||
figuring out volumes of wells so I can know exactly what volume to use for each annealing reaction. | |||
<br> | |||
purifying and concentrating all successful PCR rxns using column. should also figure out if 50x loading dye works. | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Sample Read#''' | |||
| 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''' | |||
|- | |||
| Luc14 gRNA 20||0.141||0.075||1.864||140.539||2.85 | |||
|- | |||
| Luc14 gRNA 21||0.176||0.098||1.788||175.643||2.28 | |||
|- | |||
| Luc14 gRNA 22||0.198||0.106||1.869||197.974||2.02 | |||
|- | |||
| Luc14 gRNA 27||0.131||0.07||1.868||131.014||3.05 | |||
|- | |||
| Luc14 gRNA 29||0.159||0.086||1.858||159.485||2.51 | |||
|} | |||
Set up annealing reaction, 10ul total | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''''' | |||
| align="center" style="background:#f0f0f0;"|'''ul for 400ng''' | |||
| align="center" style="background:#f0f0f0;"|'''ul elution solution''' | |||
| align="center" style="background:#f0f0f0;"|'''ul water''' | |||
| align="center" style="background:#f0f0f0;"|'''ul 10x annealing buffer''' | |||
|- | |||
| gRNA 20||2.85||0.15||6||1 | |||
|- | |||
| gRNA 21||2.28||0.72||6||1 | |||
|- | |||
| gRNA 22||2.02||0.98||6||1 | |||
|- | |||
| gRNA 27||3.05||-0.05||6||1 | |||
|- | |||
| gRNA 29||2.51||0.49||6||1 | |||
|} | |||
<br> | |||
Added 1ul of enhancer and 1ul of nuclease to each reaction, incubate at 42 for 1 hour. | |||
<br> | |||
Ran gel 2ul of ladder, 1ul of each purified PCR reaction (lost a little of gRNA 22), and gRNA 27 unpurified reaction 2 that looked like it didn't work at all. | |||
<br> | |||
uncut controls of each gRNA purified pcr of the same volumes as annealing reactions. | |||
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|} | |} | ||
__NOTOC__ | __NOTOC__ |
Latest revision as of 00:19, 27 September 2017
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09/23/2014Running 2ul of each PCR rxn on a gel, with 3ul of water and 1ul of 6x loading dye.
Set up annealing reaction, 10ul total
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