Table 1.
Strains, plasmids, and primers used in this work
| Strain, plasmid, or primer | Description | Source or reference |
|---|---|---|
| Strains | ||
| Escherichia coli Mach1 | F− φ80 (lacZ) ΔM15 ΔlacX74 hsdR(rK− mK+) ΔrecA1398 endA1 tonA | Transgene |
| Bacillus subtilis 168 | Type strain | Bacillus Genetic Stock Center (BGSC) |
| B. subtilis WB600 | trpC2 ΔnprE ΔaprE Δepr Δbpf Δmpr ΔnprB | Bacillus Genetic Stock Center (BGSC) |
| E. coli K-12 | Type strain | Laboratory stock |
| Plasmids | ||
| pEASY-T3 | Cloning vector | Transgene |
| pEASY-T3-cotG-nanA | pEASY-T3 with fusion gene cotG-nanA | This study |
| pHP13 | Low-copy-number E. coli-B. subtilis shuttle vector | Alan D. Grossman |
| pHP13-cotG-nanA | pHP13 with fusion gene cotG-nanA | This study |
| pDG1728 | E. coli-B. subtilis shuttle vector with spectinomycin resistance gene | 7 |
| pGJ103 | High-copy-number E. coli-B. subtilis shuttle vector | 39 |
| pEB03 | High-copy-number E. coli-B. subtilis shuttle vector | This study |
| pEB03-cotG-nanA | pEB03 with fusion gene cotG-nanA | This study |
| Primersa | ||
| P1 | 5′-GCCTTTGAATTCAGTGTCCCTAGCTCCGAG-3′ | This study |
| P2b | 5′-CTATTGACTAGTTGAACCCCCACCTCCTTTGTATTTCTTTTTGACTA-3′ | This study |
| P3 | 5′-AGACTAGTATGGCAACGAATTTAC-3′ | This study |
| P4 | 5′-TCCTGCAGTCACCCGCGCTCTT-3′ | This study |
| Spc-up | 5′-TTGGGCCCGAATGGCGATTTTCGTTC-3′ | This study |
| Spc-down | 5′-TGCGCTCGAGCCAATTAGAATGAATATTTC-3′ | This study |
a
The italicized letters indicate the introduction of restriction sites.
b
The underlined letters indicate the introduction of a flexible linker at the C terminus of the cotG structural gene product.