TABLE 1.
Bacterial strains and plasmids used in this study
| Strain or plasmid | Genotype or description | Reference or source |
|---|---|---|
| E. coli | ||
| DH5α | Host strain for cloning experiments | 39 |
| DH5α(pDL20-A pDL10-B pJL300-C) | Contains the H. ducreyi cdtA, cdtB, and cdtC genes on three separate plasmids; expresses CDT activity detectable in culture supernatant fluid | This study |
| DH5α(pDL19-2 pDL16-1 pLS88) | Contains three different vectors; expresses no detectable CDT activity in culture supernatant fluid | This study |
| DH5α(pJL300) | Contains H. ducreyi 35000 cdtABC gene cluster; expresses H. ducreyi CDT activity detectable in culture supernatant fluid | 44 |
| DH5α(pJL301) | Contains H. ducreyi 35000 cdtABC gene cluster with mutated cdtA gene; expresses no detectable CDT activity in culture supernatant fluid | This study |
| DH5α(pJL302) | Contains H. ducreyi 35000 cdtABC gene cluster with mutated cdtB gene; expresses no detectable CDT activity in culture supernatant fluid | This study |
| DH5α(pJL303) | Contains H. ducreyi 35000 cdtABC gene cluster with mutated cdtC gene; expresses no detectable CDT activity in culture supernatant fluid | 44 |
| HB101 | Host strain essential for propagating plasmids carrying mutated H. ducreyi DNA inserts used in H. ducreyi mutant construction | 39 |
| H. ducreyi | ||
| 35000 | Wild-type strain isolated in Winnipeg, Manitoba, Canada; produces CDT | 15 |
| 35000.301 | Isogenic cdtA mutant with kan1 cartridge inserted into cdtA gene; does not express CDT activity detectable in culture supernatant fluid | This study |
| 35000.302 | Isogenic cdtB mutant with kan1 cartridge inserted into cdtB gene; does not express CDT activity detectable in culture supernatant fluid | This study |
| 35000.303 | Isogenic cdtC mutant with cat cartridge inserted into cdtC gene; does not express CDT activity detectable in culture supernatant fluid | 44 |
| Plasmids | ||
| pBR322 | Cloning vector, Ampr Tetr | 39 |
| pUC18K | pUC18 with kan1 gene ligated into multi-cloning site | 26 |
| pACYC184 | Cloning vector capable of replication in H. ducreyi; Chlorr, Tetr | 6 |
| pLS88 | Cloning vector capable of replication in H. ducreyi; Kanr, Smr, Sulr | 10 |
| pJL300 | pBR322 with 3-kb H. ducreyi 35000 DNA insert containing cdtABC gene cluster | 44 |
| pJL301 | pJL300 with kan1 cartridge inserted into RsrII site within cdtA gene | This study |
| pJL302 | pJL300 with kan1 cartridge inserted into MscI site within cdtB gene | This study |
| pJL303 | pJL300 with cat cartridge inserted into BseRI site within cdtC gene | 44 |
| pDL19-2 | Plasmid obtained by blunt-ending and self-ligating 3.6-kb PstI-EcoRI fragment from pBR322; vector control for pDL20-A | This study |
| pDL20-A | pDL19-2 with 1.1-kb PCR-derived PstI-EcoRI DNA fragment containing H. ducreyi cdtA gene | This study |
| pDL16-1 | Plasmid obtained by blunt-ending and self-ligating 4-kb SalI-BamHI fragment from pACYC184; vector control for pDL10-B | This study |
| pDL10-B | pDL16-1 with 1.2-kb PCR-derived SalI-BamHI DNA fragment containing H. ducreyi cdtB gene | This study |
| pJL300-C | pLS88 with 813-bp PCR-derived insert containing H. ducreyi cdtC gene | 44 |