This study describes the construction of recombinant genomes that express chloramphenicol acetyltransferase (CAT) in mammalian cells. The prototype recombinant, pSV2-cat, combines the beta-lactamase gene and origin of replication from pBR322 with a simian virus 40 (SV40) early transcription region containing CAT coding sequences. CAT activity was readily measured 48 hours after introducing pSV2-cat DNA into African green monkey kidney CV-1 cells. These recombinants provided a convenient system for monitoring foreign DNA expression in tissue culture cells. Derivatives of pSV2-cat with deletions in the SV40 promoter region were constructed, showing that deletion of one 72-base-pair repeat sequence had no significant effect on CAT synthesis, while deletion of 50 base pairs from the second repeat reduced synthesis to 11% of the wild-type level. A recombinant, pSV0-cat, was also constructed with the entire SV40 promoter region removed and a unique HindIII site substituted. The study highlights the importance of using easily measurable enzymatic activities to assess promoter function in mammalian cells. CAT was chosen because it is distinguishable from endogenous activities, does not interfere with other enzymatic activities, and allows for rapid, sensitive, and reproducible assays. The pSV2-cat vectors were found to be particularly useful for studying eucaryotic promoters due to the absence of CAT activity in mammalian and avian cells. The study also demonstrated that the CAT system is effective for comparing different promoters, measuring their strength in various cell types, and evaluating transfection protocols. The study also showed that the herpes thymidine kinase promoter and the chicken alpha2 type I collagen gene promoter can drive CAT expression in mammalian cells, with the Rous sarcoma virus 3' long terminal repeat promoter being significantly more effective. The CAT system was also used to monitor the expression of exogenous genes in stably transformed cell lines. The study concluded that the CAT system provides a rapid and accurate method for evaluating promoter function in mammalian cells.This study describes the construction of recombinant genomes that express chloramphenicol acetyltransferase (CAT) in mammalian cells. The prototype recombinant, pSV2-cat, combines the beta-lactamase gene and origin of replication from pBR322 with a simian virus 40 (SV40) early transcription region containing CAT coding sequences. CAT activity was readily measured 48 hours after introducing pSV2-cat DNA into African green monkey kidney CV-1 cells. These recombinants provided a convenient system for monitoring foreign DNA expression in tissue culture cells. Derivatives of pSV2-cat with deletions in the SV40 promoter region were constructed, showing that deletion of one 72-base-pair repeat sequence had no significant effect on CAT synthesis, while deletion of 50 base pairs from the second repeat reduced synthesis to 11% of the wild-type level. A recombinant, pSV0-cat, was also constructed with the entire SV40 promoter region removed and a unique HindIII site substituted. The study highlights the importance of using easily measurable enzymatic activities to assess promoter function in mammalian cells. CAT was chosen because it is distinguishable from endogenous activities, does not interfere with other enzymatic activities, and allows for rapid, sensitive, and reproducible assays. The pSV2-cat vectors were found to be particularly useful for studying eucaryotic promoters due to the absence of CAT activity in mammalian and avian cells. The study also demonstrated that the CAT system is effective for comparing different promoters, measuring their strength in various cell types, and evaluating transfection protocols. The study also showed that the herpes thymidine kinase promoter and the chicken alpha2 type I collagen gene promoter can drive CAT expression in mammalian cells, with the Rous sarcoma virus 3' long terminal repeat promoter being significantly more effective. The CAT system was also used to monitor the expression of exogenous genes in stably transformed cell lines. The study concluded that the CAT system provides a rapid and accurate method for evaluating promoter function in mammalian cells.