High-Throughput Targeted Repeat Element Bisulfite Sequencing (HT-TREBS).
Methods Mol Biol
; 1908: 219-228, 2019.
Article
in En
| MEDLINE
| ID: mdl-30649731
ABSTRACT
High-throughput targeted repeat element bisulfite sequencing (HT-TREBS) is designed to assay the methylation level of individual retrotransposon loci of a targeted family, in a locus-specific manner, and on a genome-wide scale. Briefly, genomic DNA is sheared and ligated to Ion Torrent A adaptors (with methylated cytosines), followed by bisulfite-conversion, and amplification with primers designed to bind the targeted retrotransposon. Since the primers carry the Ion Torrent P1 adaptor as a 5'-extension, the amplified library is ready to be size-selected and sequenced on a next-generation sequencing platform. Once sequenced, each retrotransposon is mapped to a particular genomic locus, which is achieved through ensuring at least a 10-bp overlap with flanking unique sequence, followed by the calculation of methylation levels of the mapped retrotransposon using a BiQ Analyzer HT. A complete protocol for library construction as well as the bioinformatics for HT-TREBS is described in this chapter.
Key words
Full text:
1
Collection:
01-internacional
Database:
MEDLINE
Main subject:
Sequence Analysis, DNA
/
Retroelements
/
DNA Methylation
/
High-Throughput Nucleotide Sequencing
Limits:
Humans
Language:
En
Journal:
Methods Mol Biol
Journal subject:
BIOLOGIA MOLECULAR
Year:
2019
Document type:
Article
Affiliation country:
United States