FIG. 5.
Site-directed mutagenesis of the translational signals of ORF2 on P15A10. P15A10 was amplified by PCR into two separate fragments. Fragment 1–215 was amplified by using the universal −40 primer (on pTRK391) and a primer complementary to nucleotides 194 to 215 (G→C mutation at nucleotide 204). Fragment 222–888 was generated by using a primer consisting of nucleotides 222 to 243 and the lacZ primer (on pTRK391). Nucleotides 216 to 221, containing the ATG start codon for ORF2, were replaced with a HindIII site on both PCR fragments (1–215HindIII and HindIII222–888) to allow fusion. The changes made to ORF2 are indicated above the graph. The graph represents β-Gal results of P15A10 and P15A10/mutORF2. β-Gal assays were performed at least three separate times. For each assay, time point determinations were performed in duplicate.