FIG. 5.
Site-directed mutagenesis of a 20-bp region upstream of the −33 region of PE. A map indicating the location of the −33 and −7 consensus regions and the transcriptional start site (+1) is shown at the top. Two important restriction sites are indicated, as is the ATG start of orf2c. The sequence of the 20-bp region upstream of the −33 (bp −50 to bp −70) was altered by site-directed mutagenesis to a complementary sequence that preserved the spacing and G+C% composition of the region. The wild-type sequence cloned into pGFK34 was used as the positive control. pGFK63 contained the mutated sequence cloned into same vector. GUS activity was measured in extracts from cells containing each of the two vectors. The values given are for two separate assays done on three individual isolates. Standard deviations are given in parentheses.