TABLE 2.
Plasmids used in this study
| Plasmid | Description | Reference |
|---|---|---|
| pPW12 | IncP mobilizable cosmid pLAFRI with EcoRI-BamHI-EcoRI linker, Tcr, 21.6 kbp | 13 |
| pRK293 | IncP mobilizable cloning vector, Tcr Kmr, 21.3 kbp | 12 |
| pUC8 | ColE1 cloning vector, Apr, 2.7 kbp | 50 |
| pUC19 | ColE1 cloning vector, Apr, 2.7 kbp | 54 |
| pBluescript SK(+) | ColE1 cloning vector, Apr, 3.0 kbp | 1 |
| pUC4KIXX | pUC4K-based plasmid carrying the Tn5 Kmr and Blr resistance genes, Apr Kmr Blr, 4.0 kbp | 5 |
| pUCNCO | ColE1 cloning vector, pUC8 with ACCATGG sequence inserted between BamHI and HincII sites, Apr, 2.7 kbp | This study |
| p21R1 | pPW12 with a 20-kbp insert of G. diazotrophicus SRT4 containing the lsdAB locus and partial type II secretion locus, Tcr, 42 kbp | 4 |
| pALS5 | 5,181-bp SalI-EcoRI fragment from p21R1 inserted into pUC19: carries lsdABX, Apr, 7.9 kbp | 4 |
| pALS15 | pRK293 with HindIII-linearized pALS10 carrying the SRT4 lsdA gene in a 2,339-bp BglII fragment, Apr Tcr 27.2 kbp | 4 |
| pALS30 | 7.7-kbp BamHI-partial EcoRI fragment from p21R1 inserted into pUC8, carries region upstream of lsdA, Apr, 10.4 kbp | This study |
| pALS31 | 8.7-kbp BamHI-partial EcoRI fragment from p21R1 inserted into pUC8, carries region upstream of lsdA, Apr, 11.4 kbp | This study |
| pALS32 | 4,614-bp BamHI-partial EcoRI fragment from p21R1 inserted into pUC8, carries lsdBXGO, Apr, 7.3 kbp | This study |
| pALS33 | HindIII-linearized pALS30 inserted at the HindIII site of pRK293, Apr Tcr, 31.7 kbp | This study |
| pALS34 | HindIII-linearized pALS31 inserted at the HindIII site of pRK293, Apr Tcr, 32.7 kbp | This study |
| pALS35 | BamHI-linearized pALS32 inserted at the BamHI site of pPW12, Apr Tcr, 28.9 kbp | This study |
| pALS60 | 198-bp PCR band amplified from pALS5 inserted at the SmaI site of pUC19, carries N-terminal region of lsdA, Apr, 2.9 kb | This study |
| pALS89 | pUCNCO containing a 1,491-bp NcoI-HincII fragment with the nptII-ble cassette from pUC4KIXX inserted at the EcoRI site of lsdG, Apr Kmr Blr, 5.8 kbpa | This study |
| pALS113 | 1,050-bp PCR band amplified from pALS32 inserted at the SalI-BamHI sites of pUC19, carries wild-type lsdG, Apr, 3.7 kbpa | This study |
| pALS114 | 1,050-bp PCR band amplified from pALS32 inserted at the SalI-BamHI sites of pUC19, carries mutated lsdG (C162G) Apr, 3.7 kbpa | This study |
| pALS117 | HindIII-linearized pALS113 inserted at the HindIII site of pRK293, Apr Tcr, 24.8 kbp | This study |
| pALS118 | HindIII-linearized pALS114 inserted at the HindIII site of pRK293, Apr Tcr, 24.8 kbp | This study |
| pALS119 | 3,527-bp HindIII-SphI fragment of G. diazotrophicus SRT4 genome inserted into pUC19, carries lsdEF, Apr, 6.2 kbp | This study |
| pALS127 | pALS119 with the 1.2-kbp SmaI nptII cassette from pUC4KIXX inserted at the EcoRV site of lsdF, Apr Kmr, 7.4 kbp | This study |
| pALS130 | pBluescript SK (+) containing a 7,551-bp BamHI-KpnI fragment with the nptII-ble cassette from pUC4KIXX inserted at the HindIII site of lsdO, Apr Kmr Blr, 12.1 kbpa | This study |
| pALS141 | 1,967-bp SmaI-BglII fragment from pALS5 inserted at SmaI-BamHI sites in pALS60 carries lsdA, Apr 4.8 kb | This study |
| pALS143 | 1,432-bp SacII-KpnI fragment of G. diazotrophicus SRT4 genome inserted into pBluescript SK (+), carries lsdHIJ, Apr, 4.3 kbp | This study |
| pALS152 | 3,679-bp SalI-EcoRI fragment of G. diazotrophicus SRT4 genome inserted into pBluescript SK(+), carries lsdJLMN and partial lsdD, Apr, 6.4 kbp | This study |
| pALS182 | 584-bp PCR band amplified from pALS5 digested with HindIII-NcoI plus 2,114-bp NcoI-NruI fragment from pALS141 inserted at HindIII-SmaI sites in pUCNCO, carries lsdA under control of 552-bp region upstream of lsdX, Apr, 4.8 kb | This study |
| pALS184 | HindIII-linearized pALS182 inserted at the HindIII site of pRK293, Apr Tcr, 26.6 kbp | This study |
a
See text.