TABLE 2.
Primer | Positiona | Sequence (5′-3′)b |
---|---|---|
abc2Fc | −309, −288 | +GGCAATCGCTTGTGTTTTAGCG |
abc2R | +1735, +1713 | −GTGAATCCCATGATGTTGATACC |
abc3Fd | +108, +129 | +GATTGGCCGCAATGGCCGTGGG |
abc3R | +1123, +1101 | −GGTGAGCCAAAGTGGCTTCGCC |
abc3F2 | +1349, +1372 | +GATGAACCCCTTAATTACTTGG |
abc7F | −203, −182 | +GACGAAAAGAGGGATTCGTGCG |
abc7R | −11, −32 | −CCATAAAGCAAAATTGATGCAG |
abc2Fe | −309, −288 | +GGCAATCGCTTGTGTTTTAGCG |
Isa mut | −105, −85 | −GAATTCTTTAAGCTAATTTC |
Mega primer F (OG1) (double stranded)f | −309, −85 | GGCAATCGCTTGTGTTTTAGCGAATAATTTAAATTTTTATCAGAATGAAAGCAAGGGGCCTTCGTTGACAAATCAGTGAAGAGATGCTAAATTCTAGATAATAAAAGACGAAAAGAGGGATTCGTGCGTTGAAAATTTAAGTTAATTTTTTAGAGAAACTGTATAACTTTGCCTGATTGTAACGGGTGGAGTGTGCGGATTTTCGGAAATTAGCTTAAAGAATTC |
Mega primer F (V583) (double stranded)g | −309, −85 | GGCAATCGCTTGTGTTTTAGCGAATAATTTAAATTTTTATCAGAATGAAAGCAAGGGGCCTTCGTTGACAAATCAGTGAAGAGATGCTAAATTCTAGATAATAAAAGACGAAAAGAGGGATTCGTGCGTTGAAAATTTAAGTTAATTTTTTAGAGAAACTGTATAACTTTGCCTGAATATAACGGGTGGAGTGTGCGGATTTTCGGAAATTAGCTTAAAGAATTC |
Bases relative to ATG.
+, sense primer; −, antisense primer.
Primers abc2F and abc2R were used to amplify intact lsa on a 2-kb fragment.
Primers abc3F to abc7R are intragenic primers used to confirm the lsa sequence.
Primers abc2F and lsa mut were used for site-directed mutagenesis (to generate mega primer).
Mega primer F(OG1) and abc2R were used to generate lsaV-M. Boldfaced T and G were targeted in site-directed mutagenesis.
Mega primer F(V583) and abc2R were used to generate lsaOG-M. Boldfaced A and A were targeted in site-directed mutagenesis.