EpiNext CUT&RUN Fast Kit

Product Description

• Quick and reliable recovery of target protein-enriched regions
• Input cell amount as few as 500 cells, or chromatin amount as low as 0.1 µg
• Minimized immunocapture background for sequencing data analysis with <10 million reads
• Fast, streamlined procedure in less than 3h
• Highly convenient - all required components included

Overview

There are several advanced methods available for ChIP-Seq with reduced cell numbers or increased resolution. These methods include ChIP-exo and ChIPmentation. ChIP-exo provides high-resolution mapping, but is time-consuming and requires an ample supply of input cells. While ChIPmentation uses transposase and sequencing-compatible adaptors to enable the integration of ligation during the ChIP process, it follows a traditionally slow (2 days) ChIP procedure and cannot achieve high-resolution mapping. CUT&RUN (Cleavage Under Target & Release Using Nuclease) was developed to release the captured target protein/DNA complex from limited biological materials for mapping protein-DNA interaction, and it has significantly improved mapping resolution. However, CUT&RUN needs expensive PA/Mnase fusion protein that has significant A/T sequence bias, causing the target protein-interacted DNA region profiles to be seriously affected by the level of MNase digestion. Therefore, as an improvement, we have developed a new technique: cleavage under target and recover using nuclease fast (CUT&RUN Fast) for enriching histone or TF-bound DNA. Our innovative approach combines the advantages of ChIP-exo and CUT&RUN with the fastest procedures and incorporates it into the EpiNext™ CUT&RUN Fast Kit.

Principle & Procedure

The kit contains all necessary reagents required for carrying out a successful CUT&RUN starting from mammalian cells or isolated nuclei/chromatin. In the reaction, nuclei are isolated from cells. The target protein-DNA complex is bound/captured with the CHIP-grade antibody of interest. With the use of a unique nucleic acid cleavage enzyme mix, chromatin is fragmented, and DNA sequences in both ends of the target protein/DNA complex are cleaved/removed. At the same time, the DNA sequence occupied by the target protein is unaffected. The target protein-bound DNA is then purified and eluted. The enriched DNA can be used for the analysis of protein-DNA interaction with various methods, especially with next generation sequencing. Included in the kit are a positive control antibody (anti-H3K9me3), a negative control non-immune IgG, and control chromatin, which can be used to demonstrate the efficacy of the kit and performance at the PCR/bioanalyzer step.

Performance Data

Fig 2. Target protein/DNA enrichment using the EpiNext CUT&RUN Fast Kit: 
Histone/DNA complex was captured by the control antibodies (anti-H3K9me3 and anti-RNA polymerase II, Epigentek) from 100,000 Jurkat cells. Non-immuno IgG was used as control. The enriched DNA was purified and fluorescently quantified for enrichment fold comparison.