Table 1.
Strains, plasmids, and primers used in this study
| Strain or plasmid | Relevant characteristics or sequence (5′-3′) | Reference or source |
|---|---|---|
| Strains | ||
| Escherichia coli strains | ||
| S17-1 | Conjugation strain | 19 |
| XL1-Blue | QuikChange transformation strain | |
| Xanthomonas sp. strains | ||
| 35Y | Growth on poly(cis-1,4-isoprene) latex, clearing zone formation | 23 |
| 35Y-CM | Chloramphenicol-resistant mutant of 35Y | 6 |
| 35Y-CM ΔroxA-attB (SN3727) | Chromosomal deletion of roxA, attB at former roxA site, no clearing zone formation on latex agar | This study |
| 35Y-CM ΔroxA-attB/pNH1-roxA-attP (in chromosome) (SN4230) | Expression of RoxA from rhamnose promoter; Kmr Cmr; clearing zone formation in the presence of rhamnose | This study |
| 35Y-CM ΔroxA-attB/pNH1-roxA-F317A-attP (in chromosome) (SN4784) | Expression of RoxA-F317A from rhamnose promoter; Kmr Cmr | This study |
| 35Y-CM ΔroxA-attB/pNH1-roxA-F317H-attP (in chromosome) (SN4785) | Expression of RoxA-F317H from rhamnose promoter; Kmr Cmr | This study |
| 35Y-CM ΔroxA-attB/pNH1-roxA-F317L-attP (in chromosome) (SN4786) | Expression of RoxA-F317L from rhamnose promoter; Kmr Cmr | This study |
| 35Y-CM ΔroxA-attB/pNH1-roxA-F317W-attP (in chromosome) (SN4787) | Expression of RoxA-F317W from rhamnose promoter; Kmr Cmr | This study |
| 35Y-CM ΔroxA-attB/pNH1-roxA-F317Y-attP (in chromosome) (SN4788) | Expression of RoxA-F317Y from rhamnose promoter; Kmr Cmr | This study |
| Plasmids | ||
| pIC20HE | Cloning vector | 1 |
| pSI17 | Source of PhiC31 integrase gene | 20 |
| pUC9::roxA-region | Source of roxA; Apr | 6 |
| pUC9::roxA-region-ΔroxA | Precise deletion of roxA coding sequence | This study |
| pUC9::roxA-region-ΔroxA-attB | PhiC31 attB site at position of former roxA gene | This study |
| pLO3 | sacB-containing suicide vector; Tcr | 13 |
| pLO3-Km | pLO3 with 1,108-bp aphII fragment of pBBR1MCS-2 in SmaI site; Tcr Kmr | This study |
| pLO3-Km-roxA-region | pLO3 with 3,603-bp SacI fragment including roxA gene | This study |
| pLO3-Km-roxA-region-ΔroxA | Precise deletion of roxA coding sequence | This study |
| pLO3-Km-roxA-region-ΔroxA-attB | Precise deletion of roxA coding sequence with attB at former roxA sequence | This study |
| pJOE6787.1 | pBR322 ori aphII (Kmr) mob int (PhiC31 integrase) attP | J. Altenbuchner |
| pNH1 | pJOE6787.1 with rhamnose regulation genes of p4782.1 | This study |
| pNH1-roxA | Coding sequence of roxA under rhamnose promoter control | This study |
| pNH1-roxA-attP (SN4230) | Coding sequence of roxA under rhamnose promoter control, with attP site | This study |
| pNH1-roxA-attP F317A (SN4784) | RoxA-F317A expression vector with attP site; Kmr | This study |
| pNH1-roxA-attP F317H (SN4785) | RoxA-F317H expression vector with attP site; Kmr | This study |
| pNH1-roxA-attP F317L (SN4786) | RoxA-F317L expression vector with attP site; Kmr | This study |
| pNH1-roxA-attP F317W (SN4787) | RoxA-F317W expression vector with attP site; Kmr | This study |
| pNH1-roxA-attP F317Y (SN4788) | RoxA-F317Y expression vector with attP site; Kmr | This study |
| Primers | ||
| s6549 | CACGTGATGAGTAAACTTGGTCTGACAGT | |
| s6550 | CCCCGTAGAAAAGATCAAAGGAT | |
| s6551 | GGGAATATTTCGAACCCCAGAGTCC | |
| s6552 | GGGCACGTGCAAGCGCAAAGAGAAAGC | |
| s6553 | ATCATTAATCAACCCTCAGCGGATGCCC | |
| s6554 | ATCATTAATCCCGTCTCAGCGCCTAACAGG | |
| s6555 | GGGACGTCCATCGTCCACATATCCAG | |
| s6556 | GGGACGTCCCTTGTCCAGATAGCCC | |
| 1fw | ACCGCTCGAGTGCCACCTGGGATGAATGTC | |
| 1rev | TCCGCTCGAGCGCTTGGTCGGTCATTTCG | |
| 2fw | GCTCTAGAGCTAGGGTTGGCTTCACGCCGCGTAATCGAT | |
| 2rev | GCTCTAGAGCGATTCCCCCTGTACGTCCCAACGAATGAGT | |
| 3fw | GTACAGGGGGAATCGCTCTAGAGTGCCAGGGCGTGCCCTTGGGCTCCCCG | |
| 3rev | CGCGGCGTGAAGCCAACCCTAGCCGCGCCCGGGGAGCCCAAGGGCACGCC | |
| 4fw | CGGGGTACCTTCAGGTTCATCATGCCGTTTGTG | |
| 4rev | TTCCGAGCTCCTCCACGGGGAGAGCCTGAGC | |
| 5fw | TCTCCTGCAAACTGCTTTTAC | |
| 5rev | GCGAATCTGAACTATCTCATCC |