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Old 08-04-2013, 04:55 AM   #1
khata
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Exclamation NEBNext vs Illumina TruSeq ChIP-Seq library prep

Hi, I was wondering if anyone did a comparison between ChIP-Seq data obtained through NEBnext and Illumina Truseq library prep.

We're seeing huge differences between the two kits (reproducible for two different transcription factors), in particular very low enrichment in high GC regions when using Illumina reagents. The problem is most visible at TSSs, where we see a dip in coverage.

Attaching some plots to illustrate it, ChIP1 and ChIP2 are two different transcription factors, IL is Illumina, NEB NEBnext. The venns show peak-level overlaps, the histograms GC content/peak, the lineplots mean extended read count at TSSs.
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File Type: pdf ILvsNEB-summarySlide.pdf (543.8 KB, 152 views)
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Old 08-07-2013, 08:14 AM   #2
sara_ahmed
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What was the enzyme used for library aplification? It's probably a result of the difference in the enzyme used for amplification that's giving you variable coverage of GC regions.

Sara Ahmed, PhD | Director of Sequencing | Cofactor Genomics
3141 Olive St. | St. Louis, MO 63103 | tel. (314) 531-4647
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Old 08-12-2013, 05:17 AM   #3
Jweber
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@ sara_ahmed: The polymerases used in the kits are Phusion for NEBnext and a proprietary enzyme for Illumina TruSeq.
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Old 08-21-2013, 10:50 AM   #4
Hybridizationx
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You should probably use NEBNext Hi-fidelity PCR mastermix as it is significantly better than Phusion or Illumina enzyme in our hands...also the same price as Phusion too
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Old 08-22-2013, 10:28 AM   #5
gwilkie
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We've had good results with KAPA Hifi polymerase, its supposed to have very low amplification bias. Phusion is known to have poor amplification of high GC DNA.
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Old 09-16-2014, 07:38 PM   #6
BioGenomics
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Default NEB or Illumina ?

Hi all,

so is there a consensus on using NEB or Illumina ChipSeq lib prep kits ? I assume the NEB is way cheaper anyway, but seemingly had better performance from the latest post ?

thanks

Greg
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chip-seq, gc bias, illumina, neb, truseq

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