Ultradeep bisulfite sequencing analysis of DNA methylation patterns in multiple gene promoters by 454 sequencing.

摘要: We developed a novel approach for conducting multisample, multigene, ultradeep bisulfite sequencing analysis of DNA methylation patterns in clinical samples. A massively parallel sequencing-by-synthesis method (454 sequencing) was used to directly sequence >100 PCR products single run without subcloning. showed the utility, robustness, and superiority this by analyzing 25 gene-related CpG rich regions from >40 cases primary cells, including normal peripheral blood lymphocytes, acute lymphoblastic leukemia (ALL), chronic lymphocytic (CLL), follicular lymphoma (FL), mantle cell (MCL). total 294,631 sequences generated with an average read length 131 bp. On average, >1,600 individual were each amplicon far beyond few clones (<20) typically analyzed traditional sequencing. Comprehensive at molecule level using clustering algorithms revealed differential between diseases. significant increase detected ALL FL samples compared CLL MCL. Furthermore, progressive spreading periphery toward center select islands The also allowed simultaneous genetic epigenetic data association nucleotide polymorphism present LRP1B promoter. This new generation methylome will provide digital profiles aberrant human cancers offers robust classification tumor subtypes.