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Old 06-21-2012, 01:34 PM   #13
linampli
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Location: france

Join Date: Mar 2012
Posts: 7
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hi,
to get back to the library prep, I am now finalising a procedure utilising LinDA for Illumina library prep. With the modified protocol one should get an Illumina library directly with LinDA, without using PCR cycles as is the case with NuGene. Basically I incorporate the Illumina primers in the RT and 2nd strand synthesis steps.

Quote:
Originally Posted by epignome View Post
Hi, so I think I'm managing to get it to work. After amplifying 50pg I got back 48ng which is more than I expected. Unfortunately when I try to use this DNA for qPCR validation some primer sets give my much higher ct values than others. It looks as if there is some kind of a bias introduced but it doesn't seem to be dependent on GC. Any idea what this might be?
I'm also thinking about the kit I want to use to construct the libraries. Illumina's ChIP-seq seems to be laborious. Any thoughts about the new kits like NextFLEX ChIP seq or the NuGene low input library preparation kit (ready for 1ng of DNA)?
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