Table 1.
Bacterial strains, plasmids, and primers used in this study
| Strain, plasmid, or primer | Genotype, description, or sequence (5′–3′)a | Source or referenceb |
|---|---|---|
| Strains | ||
| W168 | trpC2 | BGSC no. 1A1 |
| HB13564 | W168 uppS1 yqbO::Tn7 (VAN-resistant isolate) | Tn7 library with W168 |
| HB13647 | W168 kan-uppS+ | LFH PCR with W168 |
| HB13648 | W168 kan-uppS1 | LFH PCR with W168 |
| HB13649 | W168 kan::uppS1 amyE::Pspac(hy)-uppS (cat) | pYH003 with HB13648 |
| HB13650 | W168 amyE::Pspac(hy)-uppS+-FLAG (cat) | pYH004 with W168 |
| HB13651 | W168 amyE::Pspac(hy)-uppS1-FLAG (cat) | pYH005 with W168 |
| HB13591 | W168 sigM::tet | 43 |
| HB13592 | W168 sigX::spec | 43 |
| HB13549 | W168 sigW::mls | 60 |
| HB13654 | W168 sigM::tet kan-uppS+ | HB13591 chr DNA with HB13647 |
| HB13655 | W168 sigM::tet kan-uppS1 | HB13591 chr DNA with HB13648 |
| HB13656 | W168 sigX::spec kan-uppS+ | HB13592 chr DNA with HB13647 |
| HB13657 | W168 sigX::spec kan-uppS1 | HB13592 chr DNA with HB13648 |
| HB13658 | W168 sigW::mls kan-uppS+ | HB13549 chr DNA with HB13647 |
| HB13659 | W168 sigW::mls kan-uppS1 | HB13549 chr DNA with HB13648 |
| Plasmids | ||
| pPL82 | IPTG-inducible expression vector (amyE integration) | 42 |
| pYH003 | Pspac(hy)-uppS in pPL82 | This study |
| PYH004 | Pspac(hy)-uppS+-FLAG in pPL82 | This study |
| PYH005 | Pspac(hy)-uppS1-FLAG in pPL82 | This study |
| Primers (no. [name]) | ||
| 6011 (uppS up-F) | TGAATCAGCTGTTAAGTATGAATCAC | |
| 6012 [uppS up-R(kan)] | CCTATCACCTCAAATGGTTCGCTGTGTACATAGTTTTTCATTAAACTTCCA | |
| 6014 [uppS do-F(kan)] | CGAGCGCCTACGAGGAATTTGTATCGACTGTTGATTACATTGATTATCAGCA | |
| 6015 (uppS do-R) | TACTCAGCCTTATCTCGCCG | |
| 6019 (uppS Pspac-F) | GCGCCCCGGGAACCTTTTTGGGTGACGGAG | |
| 6020 (uppS Pspac-R) | GCGCTCTAGATTTGTTTCATGTCCACCATCC | |
| 6021 (uppS-FLAG Pspac-R) | GCGCTCTAGATTACTTATCGTCGTCATCCTTGTAATCAATTCCGCCAAACCTCC |
a
Restriction sites are underlined, and the sequences complementary to antibiotic resistance cassettes for LFH PCR are in boldface.
b
chr, chromosomal.