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Chromatin immunoprecipitation analysis of Cdk8 was performed using cross-linked chromatin from 1 x 106 HCT116 colon carcinoma cells treated with serum for 0, 15, 30, and 60 minutes. Immunoprecipitation was performed using a multiplex microplate Matrix ChIP assay (see reference for Matrix ChIP protocol: http://www.ncbi.nlm.nih.gov/pubmed/22098709) with 1.0ul/100ul well volume of a Cdk8 polyclonal antibody (Product # PA1-21780). Chromatin aliquots from ~1 x 105 cells were used per ChIP pull-down. Quantitative PCR data were done in quadruplicate using 1ul of eluted DNA in 2ul SYBR real-time PCR reactions containing primers to amplify -15kb upstream of the Egr1 gene or exon-1 of Egr1. PCR calibration curves were generated for each primer pair from a dilution series of sheared total genomic DNA. Quantitation of immunoprecipitated chromatin is presented as signal relative to the total amount of input chromatin. Results represent the mean +/- SEM for three experiments. A schematic representation of the Egr-1 locus is shown above the data where boxes represent exons (black boxes = translated regions, white boxes = untranslated regions); the zigzag line represents an intron; and the straight line represents upstream sequence. Regions amplified by Egr-1 primers are represented by black bars. Data courtesy of the Innovators Program.
|Tested species reactivity||Human, Mouse, Rat|
|Host / Isotype||Rabbit / IgG|
|Immunogen||Synthetic peptide: C-SQQPPQYSHQTHRY, corresponding to amino acids 451-464 of Human Cdk8.|
|Storage buffer||PBS, pH 7.4, with 0.2% BSA|
|Contains||0.09% sodium azide|
|Storage Conditions||4° C, do not freeze|
|Tested Applications||Dilution *|
|ChIP assay (ChIP)||1-3 ul|
|Immunofluorescence (IF)||Assay dependent|
|Immunoprecipitation (IP)||10 µg/mg of lysate|
|Western Blot (WB)||1-2 µg/ml|
* Suggested working dilutions are given as a guide only. It is recommended that the user titrate the product for use in their own experiment using appropriate negative and positive controls.
PA1-21780 detects Cdk8 in Human, Rat and Mouse samples.
PA1-21780 has been successfully used in ChIP, Immunofluorescence, Immunoprecipitation and Western Blot procedures.
PA1-21780 immunogen corresponds to Synthetic peptide: C-SQQPPQYSHQTHRY, corresponding to amino acids 451-464 of Human Cdk8..
A family of proteins designated as cdks are critical regulators of cell cycle progression. The prototype member of this family, p34 / Cdc 2, and a related protein Cdk 2, function late in the cycle while Cdk 4 and Cdk 6 are critically involved in G1 to S progression. The p54 / Cdk 8 interacts exclusively with Cyclin C. The Cdk 8 Cyclin C complex activates the kinase. Because of analogy with yeast Cdk 8, it has been proposed that the human Cdk 8 Cyclin C may interact with RNA polymerase, thus influencing the transcription mechanism(s) that lead to cell division.
For Research Use Only. Not for use in diagnostic procedures. Not for resale without express authorization.
CDK8 protein kinase; Cell division protein kinase 8; Cyclin-dependent kinase 8; Mediator complex subunit CDK8; Mediator of RNA polymerase II transcription subunit CDK8; MGC126074; MGC126075; Protein kinase K35