Pipeline for large-scale microdroplet bisulfite PCR-based sequencing allows the tracking of hepitype evolution in tumors

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Abstract

Cytosine methylation provides an epigenetic level of cellular plasticity that is important for development, differentiation and cancerogenesis. We adopted microdroplet PCR to bisulfite treated target DNA in combination with second generation sequencing to simultaneously assess DNA sequence and methylation. We show measurement of methylation status in a wide range of target sequences (total 34 kb) with an average coverage of 95% (median 100%) and good correlation to the opposite strand (rho=0.96) and to pyrosequencing (rho=0.87). Data from lymphoma and colorectal cancer samples for SNRPN (imprinted gene), FGF6 (demethylated in the cancer samples) and HS3ST2 (methylated in the cancer samples) serve as a proof of principle showing the integration of SNP data and phased DNA-methylation information into hepitypes and thus the analysis of DNA methylation phylogeny in the somatic evolution of cancer.

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Herrmann, A., Haake, A., Ammerpohl, O., Martin-Guerrero, I., Szafranski, K., Stemshorn, K., … Hampe, J. (2011). Pipeline for large-scale microdroplet bisulfite PCR-based sequencing allows the tracking of hepitype evolution in tumors. PLoS ONE, 6(7). https://doi.org/10.1371/journal.pone.0021332

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