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Old 10-27-2011, 10:57 AM   #4
Location: Massachusetts

Join Date: Feb 2009
Posts: 50

Sorry, I didn't mean the PCR step. I mentioned a QIAquick PCR purification step after ligation but before size selection on a gel.

For example, the last step of "Adaptor ligation of dA-Tailed DNA" is "Purify DNA sample on one column and elute in 30uL of sterile dH2) or elution buffer" both in NEBNext kit and Illumina Paired End Library kit.

I want to know if I can directly add ligation product to a 2% gel for size selection without this purification step.
Originally Posted by HESmith View Post
The PCR insures that your library inserts are flanked by adapters, a prerequisite for cluster formation and sequencing, and also increases the amount of library. PCR-free protocols have been used successfully, but you should quantify your library by qPCR so that only adapter-ligated molecules are measured.
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