The SimpleChIP® Stem Cell Master Regulator Assay Kit contains ChIP-formulated antibodies and SimpleChIP® primers for the analysis of Oct-4, Sox2 and Nanog binding to target genes in human cells by chromatin immunoprecipitation (ChIP). The positive control SimpleChIP® Human Oct-4 Promoter Primers are provided for detection and quantification of Oct-4 promoter enrichment, as Oct-4 is a known target gene of Oct-4, Sox2 and Nanog proteins. The negative control SimpleChIP® Human α Satellite Repeat Primers allow for determination of background levels of enrichment. Antibodies and primers are tested and optimized for parallel use with the SimpleChIP® Enzymatic Chromatin IP Kits #9002 and #9003 and SYBR® Green quantitative real-time PCR. The kit provides enough reagents for 10 ChIP assays per antibody and 250 PCR reactions per primer set.
Each antibody in the SimpleChIP® Stem Cell Master Regulator Assay Kit detects endogenous levels of its respective human protein. SimpleChIP® Human Oct-4 Promoter Primers contain a mix of forward and reverse PCR primers that are specific for amplification of a 99 base pair region of the human Oct-4 promoter. SimpleChIP® Human α Satellite Repeat Primers contain a mix of forward and reverse PCR primers that are specific for the amplification of a 182 base pair region of the human α satellite repeat element.
Embryonic stem cells are pluripotent cells derived from the inner cell mass (ICM) of the mammalian blastocyst. Pluripotent cells are capable of indefinite self-renewing expansion in culture and can differentiate into cell types of all three germ layers: endoderm, ectoderm and mesoderm. This pluripotent state is a property shared by embryonic stem (ES) cells, embryonic carcinoma and induced pluripotent stem (iPS) cells. Oct-4, Sox2 and Nanog are key transcriptional regulators that are highly expressed in pluripotent cells (1). Together they form a master transcriptional regulatory network that maintains cells in a pluripotent state (2,3). Over-expression of Oct-4 and Sox2 along with Klf4 and c-Myc can induce pluripotency in both mouse and human somatic cells, highlighting their roles as key regulators of the transcriptional network necessary for self-renewal and pluripotency (4,5). It has also been demonstrated that overexpression of Oct-4, Sox2, Nanog and Lin28 can induce pluripotency in human somatic cells (6). Chromatin immunoprecipitation (ChIP) is a powerful technique that can be used to identify Oct-4, Sox2 and Nanog target genes in a given population of pluripotent cells (2,3,7-9). In addition, ChIP can be used to characterize changes in target gene occupancy that occur during induction of iPS cells from somatic cells, or differentiation of pluripotent cells into different cell lineages.
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