Sam's Notebook » in-gel PCR http://onsnetwork.org/kubu4 University of Washington - Fishery Sciences - Roberts Lab Thu, 08 Nov 2018 21:47:12 +0000 en-US hourly 1 http://wordpress.org/?v=4.0 Gel Purification & PCR cDNA SOLiD Libraries – Abalone, Yellow Perch, Lake Trout, Herring http://onsnetwork.org/kubu4/2010/04/07/gel-purification-pcr-cdna-solid-libraries-abalone-yellow-perch-lake-trout-herring/ http://onsnetwork.org/kubu4/2010/04/07/gel-purification-pcr-cdna-solid-libraries-abalone-yellow-perch-lake-trout-herring/#comments Wed, 07 Apr 2010 17:20:07 +0000 http://onsnetwork.org/kubu4/?p=676

cDNA was gel purified according to Ambion’s Whole Transcriptome Analysis Kit. The appropriate regions (100 – 200bp) were excised and cut in to 4, 1x5mm pieces. The two “internal” pieces were transferred to individual PCR tubes. The “outer” pieces were transferred together to a 1.5mL snap cap tube and stored @ -20C.

Three images are below. The first two are the gels before excising the 100 – 200bp region of the gel. The third is the image of the SECOND gel after the specified region was excised. An image was not taken of Gel 1 after excision (whoops!).

Gel 1

 

Gel 2

NOTE: The WB sample in the gell above is actually a yellow perch sample, NOT an abalone sample!

 

Gel 2 AFTER EXCISION

NOTE: The WB sample in the gell above is actually a yellow perch sample, NOT an abalone sample!

 

In-gel PCR SOLiD Libraries – Abalone, Yellow Perch, Lake Trout, Herring

In-gel PCR was performed on the individual “internal” gel pieces that were excised, as described below from earlier today. PCR rxns/cycling were performed according to Ambion’s Whole Transcriptome Analysis Kit. PCR ran O/N. PCR master mix set up is here (bottom half of sheet).

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