Differential Transient and Long-Term Expression of DNA Sequences Introduced into T-Lymphocyte Lines

Abstract

We have used a protoplast fusion protocol to introduce the genes encoding neomycin phosphotransferase (neo) and chloramphenicol acetyltransferase (CAT) into murine and human T-lymphocyte lines. Plasmid constructs containing the neo gene under the control of the promoters from the Rous sarcoma virus long terminal repeat (RSV LTR), the SV40 early region, or the herpes simplex virus thymidine kinase gene (HSV TK) can stably transform each of three T-cell lines to G-418 resistance. The characteristic frequencies for different cell lines can differ by at least two orders of magnitude, although initial DNA uptake and transient expression are similar. In the two murine cell lines, low numbers of gene copies are retained in long-term transformants. Prior to integration, transient expression assays for cat or neo gene products reveal that the differences in intrinsic promoter strength of different constructs are further influenced by the coding sequences being transcribed. Thus, while transient expression of the neo protein is similar from both the Rous LTR and the SV40 early promoter, the Rous LTR directs synthesis of CAT protein at levels two orders of magnitude higher than those from the SV40 early promoter.

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