Sensitive mapping of recombination hotspots using sequencing-based detection of ssDNA.
Genome Res
; 22(5): 957-65, 2012 May.
Article
em En
| MEDLINE
| ID: mdl-22367190
ABSTRACT
Meiotic DNA double-stranded breaks (DSBs) initiate genetic recombination in discrete areas of the genome called recombination hotspots. DSBs can be directly mapped using chromatin immunoprecipitation followed by sequencing (ChIP-seq). Nevertheless, the genome-wide mapping of recombination hotspots in mammals is still a challenge due to the low frequency of recombination, high heterogeneity of the germ cell population, and the relatively low efficiency of ChIP. To overcome these limitations we have developed a novel method--single-stranded DNA (ssDNA) sequencing (SSDS)--that specifically detects protein-bound single-stranded DNA at DSB ends. SSDS comprises a computational framework for the specific detection of ssDNA-derived reads in a sequencing library and a new library preparation procedure for the enrichment of fragments originating from ssDNA. The use of our technique reduces the nonspecific double-stranded DNA (dsDNA) background >10-fold. Our method can be extended to other systems where the identification of ssDNA or DSBs is desired.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Recombinação Genética
/
DNA de Cadeia Simples
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Mapeamento Cromossômico
Idioma:
En
Ano de publicação:
2012
Tipo de documento:
Article
País de afiliação:
Estados Unidos