TABLE 2.
Strains, plasmids, and primers used in this study
| Strain, plasmid, or primer | Description or construction | Source or reference | Sequencea |
|---|---|---|---|
| Strains | |||
| E. coli | |||
| YB4026 | S17-1/pNPTS138ΔflgE | 17 | |
| YB8286 | α-Select/pNPTS139PilAT36C | 9 | |
| YB3087 | S17-1/pLW98 | 33 | |
| YB9221 | α-Select/pUJfljK | This study | |
| YB428 | α-Select/pUJflgE | This study | |
| YB9222 | α-Select/pUJmotB | This study | |
| Caulobacter | |||
| YB8288 (parent) | NA1000 PilAT36C | 9 | |
| LS3118 (ΔpilA) | NA1000 ΔpilA | 34 | |
| YB8306 (ΔfljK) | NA1000 PilAT36C ΔfljK (lysate made from YB8287 transduced into TPA2234) | This study | |
| YB8307 (ΔflgE) | NA1000 PilAT36C ΔflgE (plasmid from strain YB4026 electroporated into strain YB8288) | This study | |
| YB8528 (ΔmotB) | NA1000::pLW98 PilAT36C ΔmotB (lysate from strain YB8524 transduced into strain YB8288) | This study | |
| FC764 | NA1000 hfsA+ | 35 | |
| YB6375 | NA1000 hfsA+ ΔflgE | 17 | |
| TPA2234 | NA1000 ΔfljK | 18 | |
| YB7377 | NA1000 hfsA+ ΔmotB | 17 | |
| YB8287 | NA1000::pNPTS139PilAT36C hfsA+ PilAT36C (electroporated strain YB8220 with plasmid from strain YB8286) | This study | |
| YB8524 | NA1000::pLW98 ΔmotB (strain YB3087 mated with YB7377) | This study | |
| YB9228 (parent EV) | NA1000 PilAT36C pUJ142 (electroporated strain YB8288 with pUJ142) | This study | |
| YB9229 (ΔpilA EV) | NA1000 ΔpilA pUJ142 (electroporated strain LS3118 with pUJ142) | This study | |
| YB9237 (ΔflK EV) | NA1000 PilAT36C ΔfljK pUJ142 (electroporated strain YB8306 with pUJ142) | This study | |
| YB9225 (ΔflK pUJfljK) | NA1000 PilAT36C ΔfljK pUJfljK (electroporated strain YB8306 with plasmid from strain YB9221) | This study | |
| YB9238 (ΔflgE EV) | NA1000 PilAT36C ΔflgE pUJ142 (electroporated strain YB8307 with pUJ142) | This study | |
| YB9230 (ΔflgE pUJflgE) | NA1000 PilAT36C ΔflgE pUJflgE (electroporated strain YB8307 with plasmid from strain YB428) | This study | |
| YB9239 (ΔmotB EV) | NA1000::pLW98 PilAT36C ΔmotB pUJ142 (electroporated strain YB8528 with pUJ142) | This study | |
| YB9231 (ΔmotB pUJmotB) | NA1000::pLW98 PilAT36C ΔmotB pUJmotB (electroporated strain YB8528 with plasmid from strain YB9222) | This study | |
| Plasmids | |||
| pNPTS138 | Litmus 38 derivative; oriT sacB Kanr | M. R. K. Alley | |
| pNPTS139 | Litmus 39 derivative; oriT sacB Kanr | M. R. K. Alley | |
| pUJ142 | Xylose-inducible promotor; Cmr | 36 | |
| pNPTS138ΔflgE | pNPTS138 containing 480-bp fragments upstream and downstream of flgE | 17 | |
| pLW98 | Suicide vector containing marker CMS18; Kanr | 33 | |
| pNPTS139PilAT36C | pNPTS139 containing 500-bp fragments upstream and downstream of PilAT36C point mutation | 9 | |
| pUJfljK | pUJ142 containing fljK gene under control of xylose-inducible promotor | This study | |
| pUJflgE | pUJ142 containing flgE gene under control of xylose-inducible promotor | This study | |
| pUJmotB | pUJ142 containing motB gene under control of xylose-inducible promotor | This study | |
| Primers | |||
| pUJfljKF | Construction of pUJfljK | TAATAA/AAGCTTGAGCAAAATGCTCCCGGCAGCCA | |
| pUJfljKR | Construction of pUJfljK | TAATAA/GGTACCCCAGACGACTCGCCCCCGTCCC | |
| 3flgEHindIIIF | Construction of pUJflgE | TAATAA/AAGCTTAGCGCTGAACCCTCGTCCCCC | |
| 3flgEKpnIR | Construction of pUJflgE | TAATAA/GGTACCGCCCTATGATTAGCGCTTAATATTCAAGAGTTCCTCA | |
| pUJmotBF | Construction of pUJmotB | TAATAA/GAATTCCTTCATGCCGCTGCTCCTAAAAAGAAGGC | |
| pUJmotBR | Construction of pUJmotB | TAATAA/AAGCTTCACAAGAGGCCACGCGCGGAGA | |
a
Boldface indicates the 5′ tail/restriction site.