TABLE 1.
Oligonucleotide primers used to amplify the type II topoisomerase genes from S. pneumoniae for protein purification and DNA sequencing
Application | Gene | Forward primer (nucleotide positions) | Reverse primer (nucleotide positions) |
---|---|---|---|
Protein purification | gyrA | 5′-ATGCAGGATAAAAATTTACTG (bp 1–21) | 5′-CTGATTTGTATTAATAAAATTGTTTAT (158–185 bp downstream of stop codon) |
gyrB | 5′-ATGACAGAAGAAATCAAAAATCTG (bp 1–24) | 5′-CATATTTTCTAGACCAAGGGAAC (17–39 bp downstream of stop codon) | |
parC | 5′-ATGTCTAACATTCAAAACATGTC (bp 1–23) | 5′-TTATTTATCTTCAGTAACTACTTC (7–30 bp downstream of stop codon) | |
parE | 5′-ATGTCAAAAAAAGGAAATCAAT (bp 1–23) | 5′-TTAAAACACTGTCGCTTCTTC (1–22 bp downstream of stop codon) | |
Target gene amplification | gyrA | 5′-TAAAAAACTTTGTCACGAATATGCC (130–105 bp upstream of start) | 5′-AACGATACGCTCACGACCAGT (bp 750–771) |
gyrB | 5′-TGAAGGACAAACCAAGACCAAA (bp 1032–1053) | 5′-GTCCATTTCACCTAGCCCCTTATA (bp 1759–1782) | |
parC | 5′-AAACCTACTCTACATTCTTTGAAAGGAG (134–106 bp upstream of start) | 5′-CAGTTGGGTGGTCAATCATGTAAA (bp 571–594) | |
parE | 5′-ACCAAGGATAAACATGGAAGCC (bp 1026–1047) | 5′-CATTCATCTCACCAAGTCCTTTGTA (bp 1737–1762) | |
QRDR amplification | gyrA | 5′-CGTTTTAGTGGTTTAGAGGC (85–66 bp upstream of start) | 5′-GACCAACTTCACTGCATCA (bp 567–585) |
gyrB | 5′-TTCTCCGATTTCCTCATG (bp 1105–1122) | 5′-CCCGGCTGGATATATTCT (bp 1665–1682) | |
parC | 5′-CGCCCTAGATACTGTGTGA (98–80 bp upstream of start) | 5′-AAATCCCAGTCGAACCAT (bp 493–510) | |
parE | 5′-TGTGGATGGAATAGTGGC (bp 1054–1071) | 5′-ACCGAACTGTTTACGGAGT (bp 1697–1715) |