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Old 10-26-2016, 02:00 AM   #1
kobeho24
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Location: HKUST, Hong Kong

Join Date: Apr 2015
Posts: 32
Default Questions about Nextera kit/tagmentation

Hi folks,
Just wonder if anyone has an experience of using the Nextera kit or home-made tagmentation assay based on the Genome Research paper. http://genome.cshlp.org/content/earl....full.pdf+html
We are not able to get libraries as expected with home made version, in particular the fragment size is hard to manipulate according to the protocol attached in the paper. (adjust input amount, modify reaction buffer composition, etc. none of them works)
But anyway, we can get amplicon after PCR with KAPA HiFi kit.
However, when we start trying to do some ATAC-seq with home-made Tn5, any purification step between tagmentation and PCR turned out to have undetectable amplicon after PCR. Whereas simply adding 0.1%-0.2% SDS to the tagmented product prior to PCR can give us a whole bunch of DNA after all.
If anyone can give me some idea, that would be much appreciated!!!

Gary
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