From: Tools for genetic manipulation of the plant growth-promoting bacterium Azospirillum amazonense
Bacterial strains | Genotype | Reference | |
---|---|---|---|
Escherichia coli XL1-Blue | recA1, endA1, gyrA96, thi-1, hsdR17(rK-, mK+), supE44, relA1, lac, [F', proAB, lacIqZΔM15::Tn10(tetr)] | Stratagene | |
Azospirillum amazonense Y2 | wild type | EMBRAPA-RJ | |
Azospirillum amazonense delK | Y2 derivative, ΔglnK | This work | |
Plasmids | Relevant characteristics | Reference | Purpose |
pUC18 | cloning vector, ampr, lacZα | Clontech | cloning procedures |
pAAGLNK | pUC18 derivative containing the SalI genomic fragment of the glnK region | This work | glnK isolation |
pGLNBA | pUC18 derivative containing the amplification product generated from the revsf_glmBint and gln_AA_do primers | This work | glnB-glnA intergenic region and partial glnA isolation |
pRK2013 | ColE1 ori, kanr, mob, tra | [45] | helper plasmid for conjugation experiments |
pHRGFPGUS | pBBR1 ori, kanr, mob, gfpmut3 | [46] | DNA transfer evaluation, reporter vectors construction |
pPZP201BK | pVS1 ori, ColE1 ori, kanr, mob | [47] | construction of pPZPLACEYFP |
pEYFP | pUC ori, eyfp, ampr | Clontech | contruction of reporter plasmids |
pPZPLACEYFP | PvuII-EcoRI fragment from pEYFP (containing the lac promoter-eyfp gene fusion) cloned into the EcoRV-EcoRI sites of pPZP201BK | This work | DNA transfer evaluation, positive control in the reporter assay |
pK19MOBSACB | Integration vector, kanr, ColE1 replication origin, mob, sacB, lacZα | [48] | glnK mutagenesis |
pKΔK | pK19MOBSACB derivative containing the flanking regions of the glnK gene joined by Crossover PCR | This work | glnK mutagenesis |
pAATEYFP | pEYFP derivative containing the BglII-NcoI fragment of pAAGLNK cloned into its NcoI-BamHI sites | This work | construction of pHRAATEYFP |
pPBEYFP | pEYFP derivative containing the intergenic region between the bcr protein superfamily gene and the glnB gene cloned into its NcoI-BamHI sites | This work | construction of pHRPBEYFP |
pPKEYFP | pEYFP derivative containing the intergenic region between the ubiH and the glnK genes cloned into its NcoI-BamHI sites | This work | construction of pHRPKEYFP |
pHRAATEYFP | HindIII-EcoRI fragment from pAATEYFP (containing the aat promoter-eyfp gene fusion) joined with the 5.8 kb HindIII-EcoRI fragment of pHRGFPGUS | This work | promoter evaluation |
pHRLACEYFP | PvuII-EcoRI fragment from pEYFP (containing the lac promoter-eyfp gene fusion) joined with the 7.4 kb EcoRV-EcoRI fragment of pHRGFPGUS | This work | promoter evaluation |
pHRPBEYFP | HindIII-EcoRI fragment from pPBEYFP (containing the glnB promoter-eyfp gene fusion) joined with the 5.8 kb HindIII-EcoRI fragment of pHRGFPGUS | This work | promoter evaluation |
pHRPKEYFP | HindIII-EcoRI fragment from pPKEYFP (containing the glnK promoter-eyfp gene fusion) joined with the 5.8 kb HindIII-EcoRI fragment of pHRGFPGUS | This work | promoter evaluation |
pHREYFP | HindIII-EcoRI fragment from pEYFP (containing the eyfp gene) joined with the 5.8 kb HindIII-EcoRI fragment of pHRGFPGUS | This work | promoter evaluation |