Methylation, the addition of methyl groups to cytosine (C), plays an important role in the regulation of gene expression in both normal and dysfunctional cells. During bisulfite conversion and subsequent PCR amplification, unmethylated Cs are converted into thymine (T), while methylated Cs will not be converted. Sequencing of this bisulfite-treated DNA permits the detection of methylation at specific sites. Through the introduction of next-generation sequencing technologies (NGS) simultaneous analysis of methylation motifs in multiple regions provides the opportunity for hypothesis-free study of the entire methylome. Here we present a whole methylome sequencing study that compares two different bisulfite conversion methods (in solution versus in gel), utilizing the high throughput of the SOLiD™ System. Advantages and disadvantages of the two different bisulfite conversion methods for constructing sequencing libraries are discussed. Furthermore, the application of the SOLiD™ bisulfite sequencing to larger and more complex genomes is shown with preliminary in silico created bisulfite converted reads.
Whole Methylome Analysis by Ultra-Deep Sequencing Using Two-Base Encoding
C. B. Bormann Chung,V. L. Boyd,K. McKernan,Yutao Fu,Cinna K. Monighetti,H. Peckham,Melissa A Barker
Published 2010 in PLoS ONE
ABSTRACT
PUBLICATION RECORD
- Publication year
2010
- Venue
PLoS ONE
- Publication date
2010-02-22
- Fields of study
Biology, Medicine
- Identifiers
- External record
- Source metadata
Semantic Scholar, PubMed
CITATION MAP
EXTRACTION MAP
CLAIMS
CONCEPTS
- bisulfite conversion
A chemical treatment that converts unmethylated cytosines to thymine while leaving methylated cytosines unchanged for downstream detection.
Aliases: bisulfite treatment, bisulfite conversion method
- in-gel bisulfite conversion
A bisulfite conversion protocol performed with DNA embedded in gel during library preparation.
Aliases: in gel conversion
- in silico bisulfite-converted reads
Computationally generated reads that simulate the sequence changes expected after bisulfite conversion.
Aliases: in silico converted reads
- in-solution bisulfite conversion
A bisulfite conversion protocol performed with DNA in solution before library construction and sequencing.
Aliases: in solution conversion
- methylation
The addition of methyl groups to cytosine residues in DNA that is the biological target measured in this work.
Aliases: DNA methylation
- pcr amplification
The polymerase chain reaction step used after bisulfite treatment to amplify converted DNA before sequencing.
Aliases: PCR
- sequencing libraries
Prepared DNA libraries constructed for high-throughput sequencing after bisulfite conversion.
Aliases: libraries
- solid system
A high-throughput next-generation sequencing platform used here to generate bisulfite sequencing data.
Aliases: SOLiD
- two-base encoding
A SOLiD sequencing readout scheme that represents nucleotides using paired-base color information.
Aliases: two base encoding, color space encoding
- whole methylome sequencing
A sequencing approach aimed at measuring methylation patterns across the entire methylome.
Aliases: methylome sequencing
REFERENCES
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