High-resolution genome-wide cytosine methylation profiling with simultaneous copy number analysis and optimization for limited cell numbers.
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.
Oda
Mayumi
M
Department of Medicine, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY, USA.
Glass
Jacob L
JL
Thompson
Reid F
RF
Mo
Yongkai
Y
Olivier
Emmanuel N
EN
Figueroa
Maria E
ME
Selzer
Rebecca R
RR
Richmond
Todd A
TA
Zhang
Xinmin
X
Dannenberg
Luke
L
Green
Roland D
RD
Melnick
Ari
A
Hatchwell
Eli
E
Bouhassira
Eric E
EE
Verma
Amit
A
Suzuki
Masako
M
Greally
John M
JM
D002478:Cells, Cultured; D003596:Cytosine / Q000378:metabolism*; D004247:DNA / Q000032:analysis* / Q000737:chemistry; D019175:DNA Methylation; D019003:Deoxyribonuclease HpaII; D015894:Genome, Human; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*
D003596:Cytosine; D004247:DNA; D019003:Deoxyribonuclease HpaII