Table 1.
Bacterial strains, plasmids, and primers used in this study
| Strain, plasmid, or primer pair | Relevant marker(s) and/or characteristic(s) or sequence | Reference, source, or purpose |
|---|---|---|
| Bacterial strains | ||
| Agrobacterium tumefaciens | ||
| 5A | Wild type, soil isolate, AsIII oxidizing | 41 |
| 5A(ΔPaioB) | aioB promoter region deletion mutant | This study |
| 5A(rpoN::aacC3) | Genr; rpoN mutant by gentamicin cassette interruption | This study |
| 5A(rpoN::aacC3)(pCPP30-rpoN) | Genr Tetr; rpoN::aacC3 complemented strain | This study |
| 5A(PrpoN) | Kmr; 5A with pLSP-PrpoN | This study |
| 5A(PaioB) | Kmr; 5A with pLSP-PaioB | Laboratory stock |
| 5A(PphoB1) | Kmr; 5A with pLSP-PphoB1 | Laboratory stock |
| 5A(PpstS1) | Kmr; 5A with pLSP-PpstS1 | Laboratory stock |
| Escherichia coli | ||
| S17-1 | Pro− Mob+; conjugation donor | 57 |
| Top10 | High-competency cloning host | Invitrogen |
| Plasmids | ||
| pCR2.1-TOPO | PCR TA cloning vector | Invitrogen |
| pCPP30 | Broad host range; tetA | 22 |
| pJQ200sk | Genr traJ oriT sacB; suicide vector | 49 |
| pLSP-kt2lacZ | Kmr oriV; lacZ fusion vector used for lacZ fusion constructs | L. Pierson, Texas A&M University |
| pCR2.1-PaioB | pCR2.1-TOPO with PaioB region | Laboratory stock |
| pCR2.1-rpoN | pCR2.1-TOPO with rpoN region | This study |
| pJQ200sk-PaioB | pJQ200sk with deleted PaioB region | This study |
| pJQ200sk-rpoN::aacC3 | pJQ200sk with rpoN gene interrupted with gentamicin cassette | This study |
| pCPP30::rpoN | pCPP30 containing the complete rpoN gene | This study |
| pLSP-PrpoN | pLSP-kt2lacZ containing rpoN promoter region | This study |
| pLSP-PaioB | pLSP-kt2lacZ containing aioB promoter region | Laboratory stock |
| pLSP-PphoB1 | pLSP-kt2lacZ containing phoB1 promoter region | Laboratory stock |
| pLSP-PpstS1 | pLSP-kt2lacZ containing pstS1 promoter region | Laboratory stock |
| Primer pairs | ||
| rpoN-f/rpoN-r (2,124 bp) | 5′-GGCGTTCTCATCACCGACCAC-3′/5′-TTTACCAGATACACGCACACTCAT-3′ | For construction of rpoN::aacC3 |
| PaioBMu-1f/PaioBMu-1r (540 bp) | 5′-GCAGCGACGCCAGTTCCTT-3′/5′-CCCATCCACTAAACTTAAACAGTGTGGGGGTTCGGTTTTC-3′ | For deletion of aioB promoter |
| PaioBMu-2f/PaioBMu-2r (547 bp) | 5′-TGTTTAAGTTTAGTGGATGGGCTTCGACATTCAGTGGAGGAG-3′/5′-CGTTGGACAGGCGGCCGTAGATGA-3′ | For deletion of aioB promoter |
| PrpoN-f/PrpoN-r (454 bp) | 5′-GGCGTTCTCATCACCGACCAC-3′/5′-GATTGCGCTCCACCTCCTG-3′ | For construction of PrpoN::lacZ |
| B22Gm-f/B22Gm-r (471 bp) | 5′-ACAAAGTTAGGTGGCTCAA-3′/5′-TGGGTCGATATCAAAGTGC-3′ | Gentamicin cassette-specific primer |
| P4/P5 | 5′-GACGTTGCCTATCCCGATGAAGAT-3′/5′-GTTTGTTGATTGGCCAGGTGTAGG-3′ | For RT-PCR of aioBa |
| phoB1-2f/phoB1-2r (361 bp) | 5′-TGTTTAAGTTTAGTGGATGGGAGGGGCGCCGGCTATTCAA-3′/5′-CGCTCTAGAGACACCGACGACCTCCCTCAG-3′ | For RT-PCR of phoB1 |
| flgBRT-f/flgBRT-r (313 bp) | 5′-CAACATCGCCAACGCCAACACA-3′/5′-CCTCTTTACCGTCATCAGCATCAT-3′ | For RT-PCR of flgB |
| nrtART-f/nrtART-r (400 bp) | 5′-CCGGCAAGGTGACGCAGAA-3′/5′-TCCCCGGTGGTACAGGCAGTGAA-3′ | For RT-PCR of nrtA |
| glnART-f/glnART-r (356 bp) | 5′-AGGCCACTCTTCGGATTG-3′/5′-CATTGATGCCTTCGTGGTTGAT-3′ | For RT-PCR of glnA |
| glnBRT-f/glnBRT-r (227 bp) | 5′-CAAGCTCGATGAAGTGAAGG-3′/5′-CGATACGTCCGGTCTGTGC-3′ | For RT-PCR of glnB |
| dctART-f/dctART-r (359 bp) | 5′-CGACAAGGCCCATGAGCAGACCA-3′/5′-CAGCGCCGAGGACCACGAACAC-3′ | For RT-PCR of dctA |
a
See reference 28.