Discover Epigenomic Variation w/ Targeted Bisulfite Sequencing | Roche Sequencing

Epigenetic researchers face a number of challenges in methylation studies related to breadth, depth, throughput and more; the SeqCap Epi Enrichment System will enable you to overcome these challenges.
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- - - - Transcript - - - -
My name is Andrew Juurinen and I’ll be your XTalks host for today. Now we’ll hear from Dr. Patrick Eimerman, International Product Manager for Target Enrichment at Roche NimbleGen. Patrick, the floor is yours.

Patrick: Thank you, Andrew. Welcome everyone to today’s webinar titled “Discover Epigenomic Variation with a Flexible Targeted Bisulfite Sequencing Method” from Roche NimbleGen.

Today I'm going to present a fairly technical overview of the NimbleGen SeqCap Epi Enrichment system which became commercially available earlier this year. The "Epi" stands for epigenetics and the subtitle for my talk is, Ultra-high complexity oligonucleotide probe pools for targeted enrichment based bisulfite sequencing: A new technology for DNA methylation analysis.

So I'll begin with a brief orientation to DNA methylation and bisulfite sequencing, then I'll spend most of my time discussing details of the SeqCap Epi technology. Finally I'll close by showing some new data from our fixed content design that Patrick mentioned.

So,DNA methylation and bisulfite sequencing. Although many methods are now available for studying DNA methylation, bisulfite mutagenesisis widely considered the "Gold" standard for single based resolution studies utilizing next generation sequencing approaches.

Bisulfite mutagenesis acts through the deamination and conversion of unmethlyated cytosines to uracils. Methlyated and hydroxymethylated cytosines are protected from this reaction and aren't converted.

After PCR amplification of bisulfite converted DNA, the uracils are replaced with thymines which are detected by DNA sequencing and in comparison to a reference sequence. Despite the power of this approach, bisulfite mutagenesis is not without problems.

The process itself damages up to 95% of DNA library fragments during the conversion. This can have serious implications for down-stream applications. However, the SeqCap Epi workflow that I'm going to talk about was designed intentionally to mitigate as much of this negative effect as possible. Initial application of bisulfite mutagenesis in the context of next-gen sequencing was for whole genome bisulfite sequencing. The most prominent early study to utilize this application was published in 2009 by Lister and colleagues.

To generate an average read depth of 14-15x per strand for two different samples which equates to approximately 90 Gb pairs of data per sample. Nearly 400 total lanes of Illumina GA2 sequencing were required. With today's sequencing technology this equates to approximately 3 lanes of HiSeq 2000 run or a HiSeq2500 run in high output mode using 2x100 reads. So, while sequencing throughput has increased dramatically, I think this is obviously still not a practical solution for most research needs.

This is where targeted enrichment or sequence capture plays its most important role. Sequence capture for enrichment from standard genomic DNA samples was first described in 2006. But the development of an effective technique for DNA methylation analysis has faced several additional challenges.

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