TABLE 1.
Bacterial strains, yeast strains, and plasmids used in this study
| Strain or plasmid | Relevant characteristics and description | Source or reference |
|---|---|---|
| A. vinelandii OP | Wild-type, nitrogen-fixing, soil bacterium | Laboratory stock |
| Escherichia coli TG1 | K-12 Δ(lac-pro) supE thi hsd-5/F′ traD36 proA+B+ lacIq lacZΔM15 | Amersham Life Sciences, Inc. |
| E. coli INVαF′ | F′ endA1 recA1 hsdR17(rK− mK+) Δ(lac)U169 φ80lacΔM15 Δ(lacZYA-argF)U169 λ− supE44 thi-1 relA1 gyrA96 | Invitrogen Corp. |
| S. cerevisiae CG1945 | MATa ura3-52 his3-200 lys2-801 ade2-101 leu2-3 trp1-901 112 gal4-542 gal180-538 cyhr2 LYS2::GAL1UAS-GAL1TATA-HIS3 URA3::GAL417-mers(×3)-CYC1TATA-lacZ | CLONTECH Laboratories, Inc. |
| pVA3-1 | Murine p53(72–390) in pAS2-1, TRP1, Ampr (9.4 kb) | CLONTECH Laboratories, Inc. |
| pTD1-1 | SV40 large T antigen (84–708) in pACT2, LEU2, Ampr (9.9 kb) | CLONTECH Laboratories, Inc. |
| pCR 2.1 | Ampr Kanr (3.9 kb); used for direct cloning of PCR products | Invitrogen Corp. |
| pBG500 | Derivative of pCR 2.1 in which 815-bp DNA fragment that encodes the central domain (catalytic domain, Fig. 1b) of NifA was cloned; this fragment was generated by PCR amplification using the primers described in the text; this fragment could be released by digesting with EcoRI and could be cloned into the EcoRI site of pAS2-1 or pGAD424 to generate in-frame fusions with GAL4 DNA BD or AD, respectively | This work |
| pBG501 | Derivative of pCR 2.1 in which 789-bp DNA fragment that encodes the central domain (catalytic domain, Fig. 1a) of the NifL was cloned; this fragment was generated by PCR amplification using the primers described in the text; this fragment could be released by digesting with EcoRI and could be cloned into the EcoRI site of pAS2-1 or pGAD424 to generate in-frame fusions with GAL4 DNA BD or AD, respectively | This work |
| pAS2-1 | GAL4(1–147) DNA-BD TRP1 AmprCYHs2 (8.5 kb); unique cloning sites include NdeI, NcoI, SfiI, EcoRI, XmaI/SmaI, BamHI, SalI, and PstI | CLONTECH Laboratories, Inc. |
| pGAD424 | GAL4(768–881) AD LEU2 Ampr hemagglutinin epitope tag (8.1 kb); unique cloning sites include SfiI, NcoI, XmaI/SmaI, BamHI, EcoRI, SacI, and XhoI | CLONTECH Laboratories, Inc. |
| pBG502 | pAS2-1 derivative in which the EcoRI fragment that encodes the catalytic domain of the NifA (obtained by EcoRI digestion of pBG500) was cloned to generate the in-frame GAL4 BD-NifA translation fusion | This work |
| pBG503 | pAS2-1 derivative in which the EcoRI fragment that encodes the kinase-like domain of the NifL (obtained by EcoRI digestion of pBG501) was cloned to generate the in-frame GAL4 BD-NifL translation fusion | This work |
| pBG504 | pGAD424 derivative in which the EcoRI fragment that encodes the catalytic domain of the NifA (obtained by EcoRI digestion of pBG500) was cloned to generate the in-frame GAL4 AD-NifA translation fusion | This work |
| pBG505 | pGAD424 derivative in which the EcoRI fragment that encodes the kinase-like domain of the NifL (obtained by EcoRI digestion of pBG501) was cloned to generate the in-frame GAL4 AD-NifL translation fusion | This work |