NEBNext ChIP-Seq library prep kit vs Illumina TruSeq kit?

ASL · 04-09-2013, 12:59 PM

Has anyone used the NEBNext ChIP-Seq Library Prep Master Mix Set (or Reagent Set) for Illumina and gotten good results?

I bought this kit on someone else's recommendation, and I'm having my samples sequenced on an Illumina MiSeq in our sequencing core, but the results have been somewhat disappointing: the read number seems to be either lower than expected, or looks very flat (not a lot of the expected peaks). Has anyone else successfully used this kit? Has anyone compared it to the Illumina TruSeq kit?

My source material is PBMCs, the ChIP was done with ENCODE-validated H3K9me3 Ab, and was enriched 22x over the input control. The library itself had a yield of ~45 ng/ul in 15 ul, and was enriched about 400x over the ChIP source material, as validated by real-time PCR, and had an insert size of 274 bp according to the Bioanalyzer chip, so it appeared to be enriched and of the correct size.....

Thanks,

weigrc · 04-10-2013, 08:35 PM

We have the experience with the NEBNext ChIP kit, and the analyzed result is quite ok.

I am afraid that your pulled-down DNAs have a lots of flank sequences to lower the number of expected tags, since the size of pulled-down DNAs we observed are less than 100 bp.

Thanks,
Wei

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04-09-2013, 12:59 PM	#1
ASL Junior Member Location: Virginia, USA Join Date: Apr 2013 Posts: 5	NEBNext ChIP-Seq library prep kit vs Illumina TruSeq kit? Has anyone used the NEBNext ChIP-Seq Library Prep Master Mix Set (or Reagent Set) for Illumina and gotten good results? I bought this kit on someone else's recommendation, and I'm having my samples sequenced on an Illumina MiSeq in our sequencing core, but the results have been somewhat disappointing: the read number seems to be either lower than expected, or looks very flat (not a lot of the expected peaks). Has anyone else successfully used this kit? Has anyone compared it to the Illumina TruSeq kit? My source material is PBMCs, the ChIP was done with ENCODE-validated H3K9me3 Ab, and was enriched 22x over the input control. The library itself had a yield of ~45 ng/ul in 15 ul, and was enriched about 400x over the ChIP source material, as validated by real-time PCR, and had an insert size of 274 bp according to the Bioanalyzer chip, so it appeared to be enriched and of the correct size..... Thanks,

04-10-2013, 08:35 PM	#2
weigrc Member Location: Hong Kong Join Date: Oct 2011 Posts: 46	We have the experience with the NEBNext ChIP kit, and the analyzed result is quite ok. I am afraid that your pulled-down DNAs have a lots of flank sequences to lower the number of expected tags, since the size of pulled-down DNAs we observed are less than 100 bp. Thanks, Wei