Hi!
We are planning to assess DNA methylation profile of 100-200 cells using bisulfite sequencing on the NextSeq500 instrument. However, the kits designed for single cells (Accel-NGS® Methyl-Seq DNA Library, Pico Methyl-Seq™ Library Prep Kit, QIAseq™ Ultralow Input Library Kit) produce genome-sized libraries that are better to sequence using more high-throughput platforms such as HiSeq or NovoSeq. Other solutions, e.g. TruSeq® Methyl Capture EPIC Library Prep Kit, generate enriched (not so big) libraries that can be sequenced using NextSeq but require a lot of input DNA.
Does anybody know solutions for methyl-seq library preparation and targeted enrichment using a low number of cells? Other words, we need a kit that is similar to TruSeq® Methyl Capture EPIC Library Prep Kit but is designed for a few cells.
Thank you in advance for your suggestions and opinions!
Best wishes,
Evgeny
Hi Evgeny,
yours is a difficult request, though one I heard many times in my 5 years working in a TCGA focused NGS Core.
I would suggest posing your question directly to your local reps at Illumina, and Zymo, along with any other biotech currently offering Bisulfite Sequencing Library Prep kits. I would hope at least ONE of them is working on a solution to this issue by now ...
Best of luck! I hope you find what you need!
Selene
Dear Selene,
Thank you for your support! We contacted Illumina but any good reply has not been received. I'll try to find any company which provides bisulfite sequencing services. Maybe, you know any of them?
Another variant is to prepare genome libraries using, for example, Pico Methyl-Seq™ Library Prep Kit and to find more high-throughput sequencher where we could sequence them.
Best,
Evgeny
Dear Dibakar,
Thanks for the links!
But according to the first paper, we need to input 160-500 ng DNA. Such amount is impossible to obtain from the microdissected samples. We could use WGA to amplify DNA, but it should be performed after bisulfite conversion. In this paper, bisulfute treating is performed after library preparation.
The second paper compares the different methods of bisulfite treating. It is beyond our problem, because we already chose the appropriate method.
Best,
Evgeny
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