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Table 1 The strains and plasmids utilized in this work

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