Home › Genetics / Genomics › Discovering CsgD Regulatory Targets in Salmonella Biofilm Using Chromatin Immunoprecipitation and High-Throughput Sequencing (ChIP-seq)
Genetics / Genomics JoVE (Open Access) Citable · DOI

Discovering CsgD Regulatory Targets in Salmonella Biofilm Using Chromatin Immunoprecipitation and High-Throughput Sequencing (ChIP-seq)

DOI: 10.3791/60736-v
What you'll learn
  • ✓ Perform chromatin immunoprecipitation (ChIP) on bacterial biofilm samples
  • ✓ Prepare and sequence ChIP DNA libraries for high-throughput sequencing
  • ✓ Identify transcription factor binding sites using ChIP-seq analysis
  • ✓ Interpret genomic interactions between CsgD and regulatory targets
Protocol

Chromatin immunoprecipitation coupled with next-generation sequencing (ChIP-seq) is a method used to establish interactions between transcription factors and the genomic sequences they control. This protocol outlines techniques for performing ChIP-seq with bacterial biofilms, using Salmonella enterica serovar Typhimurium bacterial biofilm as an example.

Difficulty
advanced
Total time
~3–5 days (biofilm growth, ChIP protocol, library prep, sequencing run)
Model organism
Salmonella enterica serovar Typhimurium
Biosafety
BSL-2

Steps

1
Perform chromatin immunoprecipitation on biofilm cells

Cross-link biofilm samples with formaldehyde, lyse cells, and immunoprecipitate CsgD-bound DNA using specific antibodies. Reverse cross-links and purify DNA for downstream analysis.

▶ 02:01
2
Prepare ChIP DNA libraries and sequence samples

Generate sequencing libraries from immunoprecipitated DNA fragments using standard protocols. Submit prepared libraries for high-throughput sequencing to identify CsgD binding sites.

▶ 12:04
3
Analyze and interpret ChIP-seq genomic results

Align sequencing reads to the Salmonella genome and identify enriched peaks representing CsgD regulatory targets. Map transcription factor interactions to biofilm-related genes.

▶ 12:41
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