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. Author manuscript; available in PMC: 2017 Apr 18.
Published in final edited form as: Cell Rep. 2016 Apr 7;15(3):599–610. doi: 10.1016/j.celrep.2016.03.038

Figure 4. PSA eRNA binds to P-TEFb.

Figure 4

(A) C4-2 cell lysate was incubated with in vitro transcribed biotin-labeled RNAs followed by western blots. Input RNA was analyzed by dot blot hybridization. Asterisk, non-specific band.

(B) C4-2 cell lysate was incubated with IgG, CDK12, FCP1, NELF-E or CYCLIN T1 antibodies for RIP, and RIP RNAs were analyzed by RT-qPCR. Data shown as means±SD (n=3). * P < 0.05.

(C) Left, sliver staining of SFB-CDK9 immunoprecipitated from 293T cells and western blot (WB). # nonspecific bands. Middle, Coomassie blue staining of GST-Pol II-C terminal domain (asterisk) used for kinase assay. Right, in vitro CDK9 kinase assay followed by WB.

(D) Top, schematic diagram of the PSA eRNA peak region and primers for RIP assay. Bottom, sonicated C4-2 cell lysate was subjected to RIP with IgG or CYCLIN T1 antibodies and RT-qPCR. * P < 0.05.

(E) ChIP assay using IgG or CYCLIN T1 antibodies or ChIRP assay using biotin-labeled LacZor PSA eRNA-specific DNA probes and streptavidin beads in C4-2 cells. Real-time PCR datashown as means±SD (n=3). * P < 0.05.