Preparation of Low-Input and Ligation-Free ChIP-seq Libraries Using Template-Switching Technology

Area of Science:

• Molecular Biology
• Genomics
• Epigenetics

Background:

• Chromatin immunoprecipitation sequencing (ChIP-seq) is crucial for genome-wide mapping of proteins.
• Low cell numbers or low-abundance targets yield insufficient DNA for standard library preparation.
• Existing methods face challenges with adapter ligation for small DNA quantities.

Purpose of the Study:

• To describe a robust ChIP-seq workflow for small cell numbers.
• To introduce a ligation-free library preparation method for sub-nanogram ChIP DNA.
• To enable efficient sequencing library preparation from limited biological samples.

Main Methods:

• A ChIP protocol for selective enrichment of DNA-binding proteins and cross-linked DNA fragments.
• A protocol for rapid cross-linking reversal and DNA recovery.
• A ligation-free library preparation using DNA SMART technology for Illumina sequencing.

Main Results:

• Successful preparation of sequencing libraries from sub-nanogram amounts of ChIP DNA.
• Demonstration of a robust, single-tube, ligation-free workflow.
• Generation of samples ready for Illumina sequencing from limited cell inputs.

Conclusions:

• The described ChIP-seq workflow effectively addresses DNA quantity limitations.
• The ligation-free library preparation method simplifies and enhances ChIP-seq for low-input samples.
• This approach expands the applicability of ChIP-seq to challenging biological contexts.