EpiNext In Situ Accessible Chromatin Sequencing (ISAC-seq) Kit

The EpiNext™ In Situ Accessible Chromatin Sequencing (ISAC-seq) Kit is a complete set of optimized reagents designed for conducting a genome-wide analysis of chromatin accessibility, including open/closed chromatin profiling and nucleosome/transcription factor positioning from biological samples via next generation sequencing. The kit has the following advantages and features:

• Extremely fast three-step protocol: Complete the procedure (from cell sample to sequencing-ready library DNA) in just 1 hour and 35 minutes, much faster than ATAC-seq (4 hours). Minimizes nuclear damage by allowing the assay to be in in situ status, thereby preserving chromatin structure.
• Versatile usage: Works with small amounts of fresh/frozen cells, even in harsh environments, unlike DNAse-seq/MNase-seq.
• No Transposase Bias: Absence of sequence-specific cleavage minimizes fragmentation bias and biases related to GC or AT content.
• Higher accessible chromatin site coverage: Sequence-independent binding allows researchers to achieve higher accessible chromatin site coverage, unlike ATAC-seq, which shows reduced coverage at less accessible chromatin sites due to sequence-specific binding.
• Comprehensive kit: Includes all necessary components for each step, ensuring convenience and cost-effectiveness of the EpiNext™ In Situ Accessible Chromatin Sequencing (ISAC-seq) Kit.
  
• No Special Software: Use existing, time-tested ChIP-seq bioinformatics pipelines for NGS data analysis (no special ATAC-seq settings are needed due to no transposase bias).
  

Background Information
Determination of chromatin accessibility through identifying open/closed chromatin and mapping the nucleosome positioning along the genome is directly linked to epigenetic gene regulation and chromatin states and structure. Several methods are currently used to detect chromatin accessibility. The traditional method involves employing DNase I (DNase-seq) or micrococcal nuclease (MNase-seq) as tools to assess chromatin accessibility. However, these methods may have drawbacks that limit them from broad-spectrum use. They require a significant amount of starting material and are susceptible to assay bias due to challenges regulating enzyme concentrations and digestion duration. Moreover, these methods need to use isolated chromatin, which may not preserve native chromatin structure and spatial organization of regulatory elements within a cell, thereby potentially disrupting important chromatin interactions and altering accessibility patterns. The assay for transposase-accessible chromatin with next generation sequencing (ATAC-seq) is a technique that was first described as an advanced alternative method to MNase-seq and DNase-seq. In ATAC-seq, intact nuclei are directly used, and accessible DNA regions are identified by probing open chromatin with hyperactive mutant Tn5 transposase that inserts sequencing adapters into open chromatin regions in an in situ status. This method is shown to be faster and more sensitive in epigenome analysis than DNase-seq or MNase-seq and needs much less starting material amount. However, ATAC-seq has significant fragmentation and GC content bias because of sequence-specific cleavage of the Tn5 enzyme. Additionally, it is disadvantaged by covering less accessible chromatin sites compared to MNase-seq and DNase-seq.

Principle & Procedure
This kit contains all the necessary reagents required to obtain a genome-wide analysis of chromatin accessibility from cell/tissue samples via NGS. In this assay, cells are directly exposed to a unique nucleic acid cleavage enzyme mix after permeabilization. Chromatin is fragmented, and DNA sequences at both ends of the target chromatin are cleaved/removed. At the same time, the DNA sequence occupied by the protein/histone is unaffected. The adaptors are ligated to the protein/histone-bound DNA fragments. The ligated DNA is then released, purified, and amplified with a high-fidelity PCR mix for library DNA construction.

Starting Materials
Starting materials can include various cell samples such as culture cells from a flask or plate, primary cells, or rare cell populations isolated from blood, bodily fluid, fresh/frozen tissues, specific cells sorted from entire cell populations and embryonic cells, etc.

Datasheet/User Guide If the above file is not available online, you can request a copy by contacting us.