cDNAs encoding poly(ADP-ribose) polymerase from a human hepatoma lambda gt11 cDNA library were isolated by immunological screening. One insert of 1.3 kilobases (kb) consistently hybridized on RNA gel blots to an mRNA species of 3.6-3.7 kb, which is consistent with the size of RNA necessary to code for the polymerase protein (116 kDa). This insert was subsequently used in both in vitro hybrid selection and hybrid-arrested translation studies. An mRNA species from HeLa cells of 3.6-3.7 kb was selected that was translated into a 116-kDa protein, which was selectively immunoprecipitated with anti-poly (ADP-ribose) polymerase. To confirm that the 1.3-kb insert from lambda gt11 encodes for poly(ADP-ribose) polymerase, the insert was used to screen a 3- to 4-kb subset of a transformed human fibroblast cDNA library in the Okayama-Berg vector. One of these vectors [pcD-p(ADPR)P; 3.6 kb] was tested in transient transfection experiments in COS cells. This cDNA insert contained the complete coding sequence for polymerase as indicated by the following criteria: A 3-fold increase in in vitro activity was noted in extracts from transfected cells compared to mock or pSV2-CAT transfected cells. A 6-fold increase in polymerase activity in pcD-p(ADPR)P transfected cell extracts compared to controls was observed by "activity gel" analysis on gels of electrophoretically separated proteins at 116 kDa. A 10- to 15-fold increase in newly synthesized polymerase was detected by immunoprecipitation of labeled transfected cell extracts. Using pcD-p(ADPR)P as probe, it was observed that the level of poly(ADP-ribose) polymerase mRNA was elevated at 5 and 7 hr of S phase of the HeLa cell cycle, but was unaltered when artificial DNA strand breaks are introduced in HeLa cells by alkylating agents.

Alkhatib, H. M., Chen, D. F., Cherney, B., Bhatia, K., Notario, V., Giri, C., Stein, G., Slattery, E., Roeder, R. G., & Smulson, M. E. (1987). Cloning and expression of cDNA for human poly(ADP-ribose) polymerase. Proceedings of the National Academy of Sciences, 84(5), 1224–1228.