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High-resolution genome-wide cytosine methylation profiling with simultaneous copy number analysis and optimization for limited cell numbers.
Nucleic Acids Res ; 37(12): 3829-39, 2009 Jul.
Article em En | MEDLINE | ID: mdl-19386619
Many genome-wide assays involve the generation of a subset (or representation) of the genome following restriction enzyme digestion. The use of enzymes sensitive to cytosine methylation allows high-throughput analysis of this epigenetic regulatory process. We show that the use of a dual-adapter approach allows us to generate genomic representations that includes fragments of <200 bp in size, previously not possible when using the standard approach of using a single adapter. By expanding the representation to smaller fragments using HpaII or MspI, we increase the representation by these isoschizomers to more than 1.32 million loci in the human genome, representing 98.5% of CpG islands and 91.1% of refSeq promoters. This advance allows the development of a new, high-resolution version of our HpaII-tiny fragment Enrichment by Ligation-mediated PCR (HELP) assay to study cytosine methylation. We also show that the MspI representation generates information about copy-number variation, that the assay can be used on as little as 10 ng of DNA and that massively parallel sequencing can be used as an alternative to microarrays to read the output of the assay, making this a powerful discovery platform for studies of genomic and epigenomic abnormalities.
Assuntos

Texto completo: 1 Base de dados: MEDLINE Assunto principal: DNA / Reação em Cadeia da Polimerase / Metilação de DNA / Citosina Limite: Humans Idioma: En Ano de publicação: 2009 Tipo de documento: Article

Texto completo: 1 Base de dados: MEDLINE Assunto principal: DNA / Reação em Cadeia da Polimerase / Metilação de DNA / Citosina Limite: Humans Idioma: En Ano de publicação: 2009 Tipo de documento: Article