Haynes Lab:Notebook/RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines/2015/04/15: Difference between revisions
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|style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines</span> | |style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]]<span style="font-size:22px;">RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines</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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==Plasmid Maxiprep of KAH187-MV8== | ==Plasmid Maxiprep of KAH187-MV8== | ||
<b>--Notes--</b> | |||
During the plasmid purification procedure, I accidentally mixed the lysate flowthrough (containing plasmid) with the column preparation solution that was run through the Binding Column. Proceeding as normal from this point forward. Hopefully doesn't influence yield. | |||
----- | |||
<b>--DNA Quantification--</b> | |||
Measured using the Take3 plate: | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Sample ID''' | |||
| align="center" style="background:#f0f0f0;"|'''Concentration (ng/µL)''' | |||
| align="center" style="background:#f0f0f0;"|'''St Dev (ng/µL)''' | |||
|- | |||
| KAH187-MV8||163.5||7.1 | |||
|} | |||
----- | |||
<b>--Restriction Enzyme Digest--</b> | |||
{| {{table}} | |||
| align="center" style="background:#f0f0f0;"|'''Reagent''' | |||
| align="center" style="background:#f0f0f0;"|'''Volume''' | |||
|- | |||
| DNA||600ng (4uL) | |||
|- | |||
| 10x FastDigest Buffer||3 | |||
|- | |||
| EcoRI||1 | |||
|- | |||
| ApaLI||1 | |||
|- | |||
| Water||21 | |||
|- | |||
| Total||30 | |||
|} | |||
Digest for 15 minutes at 37°C. | |||
----- | |||
<b>--Gel Electrophoresis--</b> | |||
[[Image:2015-04-15_KAH187-MV8_digest_annotated.png]] | |||
Expected size fragments from EcoRI/ApaLI digestion: 3951, 1243, 1135, 288, 234. | |||
Gel was run a little too long, ~300bp fragment sizes are difficult to visualize. | |||
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Latest revision as of 00:54, 27 September 2017
 RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines
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Plasmid Maxiprep of KAH187-MV8--Notes-- During the plasmid purification procedure, I accidentally mixed the lysate flowthrough (containing plasmid) with the column preparation solution that was run through the Binding Column. Proceeding as normal from this point forward. Hopefully doesn't influence yield. --DNA Quantification-- Measured using the Take3 plate:
--Restriction Enzyme Digest--
Digest for 15 minutes at 37°C. --Gel Electrophoresis--
Expected size fragments from EcoRI/ApaLI digestion: 3951, 1243, 1135, 288, 234. Gel was run a little too long, ~300bp fragment sizes are difficult to visualize.
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