Chapter title |
Genome-Wide Profiling of DNA Methyltransferases in Mammalian Cells
|
---|---|
Chapter number | 9 |
Book title |
CpG Islands
|
Published in |
Methods in molecular biology, January 2018
|
DOI | 10.1007/978-1-4939-7768-0_9 |
Pubmed ID | |
Book ISBNs |
978-1-4939-7767-3, 978-1-4939-7768-0
|
Authors |
Massimiliano Manzo, Christina Ambrosi, Tuncay Baubec |
Abstract |
Chromatin immunoprecipitation followed by high-throughput sequencing (ChIP-seq) is currently the method of choice to determine binding sites of chromatin-associated factors in a genome-wide manner. Here, we describe a method to investigate the binding preferences of mammalian DNA methyltransferases (DNMT) based on ChIP-seq using biotin-tagging. Stringent ChIP of DNMT proteins based on the strong interaction between biotin and avidin circumvents limitations arising from low antibody specificity and ensures reproducible enrichment. DNMT-bound DNA fragments are ligated to sequencing adaptors, amplified and sequenced on a high-throughput sequencing instrument. Bioinformatic analysis gives valuable information about the binding preferences of DNMTs genome-wide and around promoter regions. This method is unconventional due to the use of genetically engineered cells; however, it allows specific and reliable determination of DNMT binding. |
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