An Integrated Workflow to Study the Promoter-Centric Spatio-Temporal Genome Architecture in Scarce Cell Populations

J Vis Exp. 2023 Apr 21:(194). doi: 10.3791/65316.

Abstract

Spatiotemporal gene transcription is tightly regulated by distal regulatory elements, such as enhancers and silencers, which rely on physical proximity with their target gene promoters to control transcription. Although these regulatory elements are easy to identify, their target genes are difficult to predict, since most of them are cell-type specific and may be separated by hundreds of kilobases in the linear genome sequence, skipping over other non-target genes. For several years, Promoter Capture Hi-C (PCHi-C) has been the gold standard for the association of distal regulatory elements to their target genes. However, PCHi-C relies on the availability of millions of cells, prohibiting the study of rare cell populations such as those commonly obtained from primary tissues. To overcome this limitation, low input Capture Hi-C (liCHi-C), a cost-effective and customizable method to identify the repertoire of distal regulatory elements controlling each gene of the genome, has been developed. liCHi-C relies on a similar experimental and computational framework as PCHi-C, but by employing minimal tube changes, modifying the reagent concentration and volumes, and swapping or eliminating steps, it accounts for minimal material loss during library construction. Collectively, liCHi-C enables the study of gene regulation and spatiotemporal genome organization in the context of developmental biology and cellular function.

Publication types

  • Video-Audio Media
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Chromatin
  • Enhancer Elements, Genetic
  • Gene Expression Regulation
  • Genome*
  • Promoter Regions, Genetic
  • Regulatory Sequences, Nucleic Acid*
  • Workflow

Substances

  • Chromatin