• Le 03 décembre 2018
    Institut de Recherche en Santé - 8 quai Moncousu - Nantes
    Amphithéâtre Denis Escande
  • 11h30

Biochemical analysis of chromatin mechanisms using the locus-specific chromatin immunoprecipitation technology

Biochemical analysis of chromatin mechanisms using the locus-specific chromatin immunoprecipitation technology

Hodaka FUJII, MD, PhD, invited by Karim Si-Tayeb (Eq IV)
Professor at Department of Biochemistry and Genome Biology, Hirosaki University Graduate School of Medicine
Hirosaki, Japan

ABSTRACT

Comprehensive understanding of chromatin mechanisms requires identification of molecules (proteins, RNA, genomic regions, etc.) associated with specific genomic regions of interest in vivo. To this end, we developed the locus-specific chromatin immunoprecipitation (locus-specific ChIP) technology consisting of insertional ChIP (iChIP) and engineered DNA-binding molecule-mediated ChIP (enChIP) to purify the genomic regions of interest.

In iChIP, specific genomic regions tagged with the recognition sequences of an exogenous DNA-binding protein such as LexA are subjected to affinity purification. In enChIP, specific genomic regions are tagged with engineered DNA-binding molecules such as transcription activator-like (TAL) proteins and the clustered regularly interspaced short palindromic repeats (CRISPR) system consisting of a catalytically inactive form of Cas9 (dCas9) plus guide RNA (gRNA) for biochemical purification. In addition, we have recently developed in vitro iChIP and in vitro enChIP systems using recombinant and/or synthetic DNA-binding molecules.

iChIP and enChIP are complimentary and comprehensive approaches with an emphasis on non-biased search using next-generation sequencing (NGS), microarrays, mass spectrometry (MS), and other methods. In addition, this approach is not restricted to cultured cell lines but easily extended to organisms.

We will present recent advances in the locus-specific ChIP technology including its application to identification of long-range interactions between genomic regions. The locus-specific ChIP technology would be useful for elucidation of chromatin mechanisms.