Welander and Metcalf. 10.1073/pnas.0502623102. |
Supporting Table 4
Table 4. Plasmids and primers used in the study
Plasmid or primer | Description and/or construction or primer sequence | Source, reference, or added site |
Plasmids | ||
pBluescript SK(+) | ApR cloning vector | Strategene |
pTn5-407 | pac-ori-aph cassette plasmid; KnR | 1 |
pPW3 | XbaI-SstI cut mtr downstream PCR fragment amplified using primers PW3 and PW4 and ligated into XbaI-SstI cut pBluescript SK(+) | This study |
pPW4 | XbaI-KpnI cut mtr upstream PCR fragment amplified using primers PW1 and PW4 and ligated into XbaI-KpnI cut pPW3 | This study |
pPW7 | pac-ori-aph cassette flanked by mtr upstream and downstream regions; constructed by ligating XbaI cut pTn5-407 to XbaI cut pPW4 | This study |
pPW23 | Copy control plasmid that contains a multiple cloning site flanked by the M. barkeri Fusaro hpt upstream and downstream regions | This study |
pPW25-3 | SstI-NotI cut mtr operon plus 1kb upstream and downstream PCR amplified using primers PW122 and PW123 and inserted between the hpt regions of SstI-NotI cut pPW23 | This study |
Primers | ||
PW1 | CGGAATTCGGTACCACTAGTTTCAATTTCAAGTGTCGAACCA | KpnI, SpeI |
PW2 | TTGGCGCGCCTCTAGACATATGTTTTCCTCCTTTGTATTATTACA | XbaI, NdeI |
PW3 | TTGGCGCGCCTCTAGAGGATCCGCTCTGACTACCTCCTTTACGG | XbaI, BamHI |
PW4 | CGGAATTCGAGCTCGCGGCCGCGCCAAGTCTACCCCAACAAA | SstI, NotI |
PW122 | TTGGCGCGCCGAGCTCCGAACCATTGTAAATCCCAAG | SstI |
PW123 | CGGAATTCGCGGCCGCGCCAAGTCTACCCCAACAAA | NotI |
1. Zhang, J. K., White, A. K., Kuettner, H. C., Boccazzi, P. & Metcalf, W. W. (2002) J. Bacteriol. 184, 1449-1454.