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. 1981 Oct;78(10):6386–6390. doi: 10.1073/pnas.78.10.6386

Transcription of the simian virus 40 genome in DNA-transformed murine teratocarcinoma stem cells.

A Linnenbach, K Huebner, C M Croce
PMCID: PMC349044  PMID: 6273868

Abstract

To study the molecular basis for lack of expression of the simian virus 40 (SV40) early region genes in murine teratocarcinoma-derived stem cells, we introduced a recombinant plasmid consisting of pBR322 linked to the herpes simplex virus type 1 thymidine kinase gene and SV40 genome into thymidine kinase-deficient F9 stem cells. The resulting stem cell clone, 12-1, and a retinoic acid-induced differentiated daughter cell clone, 12-1a, each contain one copy per cell of the entire recombinant plasmid integrated into the cellular genome through a site on the pBR322 genome. Restriction endonuclease analyses indicate that there is no difference in integration site or organization of the three component parts of the plasmid genome within cellular DNA of stem and differentiated cells; yet the differentiated cells, 12-1a, express SV40 large tumor antigen whereas the stem cells, 12-1, do not. Both stem and differentiated cells produce two size classes of polyadenylylated RNA, 2900 and 2600 bases in length, homologous to the early region of the SV40 genome, detectable by RNA blotting analysis. S1 nuclease analysis of the SV40 transcripts present in stem and differentiated cells indicate that the SV40 mRNAs were identically spliced in the two cell types, in a manner consistent with that observed for spliced large and small tumor antigen mRNAs in SV40-infected monkey kidney cells. Thus, the failure of 12-1 teratocarcinoma stem cells, containing an integrated SV40 genome, to express SV40 tumor antigen is not due to a lack of transcription of the SV40 early region or to an inability to splice primary transcripts.

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