The field of epigenetics has grown tremendously over the past several decades. Studies have shown that chromatin modifications, DNA methylation, and DNA hydroxymethylation play critical roles in the regulation of gene expression, gene silencing, protein-DNA interactions, and other cellular processes. Dysregulation of epigenetic modifications can contribute to developmental abnormalities, neurological disorders, and even cancer.

Chromatin immunoprecipitation (ChIP) is the prevailing method used for the study of protein-DNA interactions and the dynamics of epigenetic modifications. ChIP facilitates the identification of regions of the genome associated with a specific protein.  The Zymo-Spin^™ ChIP Kit from Zymo Research provides a streamlined ChIP procedure for investigating protein-DNA interactions that have been “fixed” in their natural state and can be used to effectively identify binding sites for transcription factors, co-factors, and other DNA regulatory proteins.

Briefly, this ChIP protocol involves covalent cross-linking of protein-DNA complexes with formaldehyde followed by cell lysis and chromatin shearing. A ChIP-grade antibody (user supplied) is used with Protein A magnetic beads to immunoprecipitate the protein-DNA complexes of interest.  Following reverse crosslinking, RNase A and Proteinase K treatments, the DNA is eluted in a minimal volume of buffer using a unique micro-elution spin column, eliminating the need for messy precipitations.  The protocol has been optimized for efficient crosslinking, shearing, and immunoprecipitation regardless of the mammalian cell type.  Additionally, eluted ChIP DNA is ideal for end point PCR, quantitative PCR, ChIP-Seq library preparation, and Next-Gen sequencing-based applications.

Equipment	Sonicator, magnetic tube rack, vortex, tube rotator, microcentrifuge, agarose gel apparatus/Bioanalyzer/TapeStation, qPCR instrument.
Compatibility	This kit contains reagents optimized for mammalian cells.
User Supplied Reagents	Formaldehyde (37% stock, w/v), Glycine (2.5 M), Phosphate Buffered Saline, Protease inhibitor cocktail, Phenylmethylsulfonyl Fluoride (0.1 M), Ethanol (95% or 100%), ChIP-grade antibody, QuestTaq™ qPCR Premix

Outline of the Zymo-Spin™ ChIP Kit Procedure

ChIP DNA Purification Method	Zymo Research	Phenol-chloroform	Supplier Q
Total Time	5 min.	>60 min.	5 min.

ChIP DNA Purification Comparison

ChIP assays were performed with HeLa cells and ChIP-grade anti-H3K4me3 and rabbit IgG. Both total and immunoprecipitated chromatin were reverse cross-linked and recovered using either the ChIP DNA Clean & Concentrator™ (included in the Zymo-Spin™ ChIP Kit), phenol-chloroform extraction, or DNA recovery kit from Supplier Q. The amount of ChIP DNA was determined using qPCR with primers specific to the GAPDH promoter. ChIP DNA enrichment is graphed as % input (i.e., the relative amount of immunoprecipitated DNA compared to input DNA following qPCR analysis).

Unique workflow features a micro-elution (≥6 µl) spin column for purification of ChIP DNA.

High quality ChIP DNA is ideal for ChIP-qPCR, ChIP-Seq, and other molecular applications.

Product	Size	Catalog #	How to Buy
Zymo-Spin™ ChIP Kit	10 preps	D5209
Zymo-Spin™ ChIP Kit	25 Preps	D5210

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