Bio-Rad Laboratories, Inc.’s EpiQ chromatin analysis kit is a real-time PCR assay for the rapid quantitative assessment of chromatin structure. Complementing existing epigenetic assays such as DNA methylation and chromatin immunoprecipitation, the EpiQ kit is the first commercial research tool that helps scientists quantify the impact of epigenetic events on gene expression regulation through chromatin state changes.
“The EpiQ chromatin analysis kit provides researchers with an easy and fast way to quantify a gene’s chromatin state,” said Viresh Patel, Bio-Rad Marketing Manager for PCR reagents. “Using the kit, cancer, developmental, and stem cell biology researchers are able to glean direct information regarding the chromatin state that’s associated with epigenetic marks such as DNA methylation and histone modifications for the first time.”
Epigenetic processes such as DNA methylation and histone modification control gene expression by altering chromatin structure. Genes that are actively transcribed are associated with “accessible” chromatin regions called euchromatin, while genes that are transcriptionally silent are often in “inaccessible” chromatin regions called heterochromatin. The EpiQ kit can provide quantitative information about chromatin accessibility, which correlates very strongly with gene expression.
Until now, the only way to quantitatively probe the status of chromatin structure was through the use of homebrew methods, first described in 2001. This approach requires the isolation and purification of nuclei from several millions cells prior to chromatin digestion and takes more than two days to complete. With the EpiQ kit, chromatin structure data can be obtained within six hours from as few as 50,000 cultured cells, without the need for nuclei isolation.
The Chromatin state of a gene can be identified in situ based on its sensitivity to the action of the nuclease in the EpiQ kit. In heterochromatin, genomic DNA is inaccessible to nuclease digestion and remains available for subsequent qPCR. Analysis of heterochromatin using the EpiQ kit reveals a minimal quantification cycle (Cq) shift between digested and undigested samples. In contrast, DNA in euchromatin is susceptible to nuclease digestion and is unavailable for qPCR. Analysis of euchromatin using the EpiQ kit shows a large Cq shift between digested and undigested samples.
EpiQ kit components include buffers for cell permeabilization and in situ chromatin digestion, optimized nuclease, materials for genomic DNA purification, control assays (qPCR primers) for chromatin assessment of a reference (epigenetically silenced) and control (constitutively expressed) gene, and EpiQ Chromatin SYBR Green Supermix, a real-time PCR reagent designed to amplify genomic DNA. The supermix can be used on all Bio-Rad real-time PCR instruments and on those from other suppliers; some instruments may require slight modifications to the supermix.