[27] Use of the CAT reporter gene for optimization of gene transfer into eukaryotic cells

1987 
Publisher Summary This chapter considers the use of the gene that confers bacterial resistance to the antibiotic chloramphenicol in a reporter gene system. The gene product is an enzyme—chloramphenicol acetyltransferase (CAT)—that is peculiar to prokaryotes. As such, no significant expression is demonstrable in eukaryotic cells that may serve as targets. Rapid and simple assays for CAT activity have been available as the investigations of bacterial resistance by Shaw. Use of an enzymatic assay for the gene product can facilitate detection of even low levels of product. Experiments in Saccharomyces cerevisiae suggested that CAT enzyme could be expressed in and are apparently nontoxic to eukaryotes. Some scientists have constructed expression vectors in which the CAT gene was fused to the SV40 early promoter (pSV2CAT) or to the 3' long terminal repeat of Rous sarcoma virus (pRSVCAT). In conclusion, the high level expression from pRSVCAT or pSV2CAT coupled with the sensitivity of the CAT enzymatic assay should continue to be of use in the optimization and development of techniques for the transfer of genes into cells.
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