FIG. 1.

Construction of a porcine LT₁₉₂:STa-toxoid genetic fusion. PCR primers 1(184EcoRV-F) and 6 (LT-R) amplified the entire porcine eltAB genes (without a stop codon), and primers 5 (STa-F) and 4 (pBREagI-R) amplified the full-length porcine estA gene (without a signal peptide). Primers 7 (LT₁₉₂-R) and 8 (LT₁₉₂-F; complementary to primer 7) mutated eltAB genes for LT₁₉₂, and primers 10 (mSTa12-R) and 9 (mSTa12-F; complementary to primer 10) mutated the STa gene to produce an STa mutant. Primers 2 (pLT:STa-R) and 3 (pLT:STa-F) added a Gly-Pro linker and genetically fused the mutated LT genes and the mutated STa gene. The gene sizes are not proportional. The lower right panels show Western blot detection of toxoid fusions using anti-CT and anti-STa antibodies, and total protein samples from TOPO cells were used as a negative control.