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Old 09-04-2015, 10:03 AM   #1
njlodato
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Location: Boston, MA

Join Date: Sep 2014
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Default HiSeq insert size

Does anyone have experience with sequencing libraries with larger-than-normal insert sizes on the HiSeq 2000/2500? Iím getting ready to prepare ChIP-Seq libraries (using NEB Next Ultra DNA Kit) and Iím leaning towards using material thatís 1kb+ in size. I know sequencing something this long is not idea for ChIP-Seq in terms of identifying binding sites, although in this case I ChIPíd for K27me3, but we are leaning towards using large material for several reasons. The most important reason is we just see better signal (% input via qPCR of known K27me3 regions) and higher signal/noise ratios (qPCR of known K27me3 regions vs qPCR of known K27me3 depleted regions) in the material greater than 1100bp after SPRI size selection than in any other fraction or size range.

Thanks for any and all input and apologies for posting about a topic that has probably been discussed many times before.
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