Optimal conditions for transformation of Azotobacter vinelandii

Page, William; Tigerstrom, Margaret von

doi:10.1128/jb.139.3.1058-1061.1979

Cited by 128 publications

(63 citation statements)

References 16 publications

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“…A. vinelandii cells were made competent according to the procedure described previously [22]. In brief, A. vinelandii was cultured on BN Fe 3 agar for about 36 h at 30³C.…”

Section: Preparation Of Competent Cells Of Azotobactermentioning

confidence: 99%

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Segregation pattern of kanamycin resistance marker in Azotobacter vinelandii did not show the constraints expected in a polyploid bacterium

Pulakat

1998

FEMS Microbiology Letters

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It was suggested that Azotobacter vinelandii cells contain about 80 copies of their chromosome and when foreign genes are introduced into the cell, it took several generations for them to spread to all 80 chromosomes even in the presence of selection. In contrast, the fact that many recessive mutants can be isolated from A. vinelandii without the constraints expected for a cell that has 80 copies of its chromosome argued against this organism being highly polyploid. We have investigated the segregation of a kanamycin resistant genetic marker under non-selective conditions in A. vinelandii. Plasmid DNA was used to introduce the kanamycin resistance gene onto the A. vinelandii chromosome at the nifY locus by homologous recombination. The transformants were identified from non-transformants with the aid of replica plating, and hence the colonies examined for segregation of the genetic marker were never subjected to kanamycin selection. In spite of growing the transformants in the absence of selection pressure, no segregant that lacked the kanamycin resistance gene was scored. These analyses suggested that the segregation of the kanamycin marker in A. vinelandii did not exhibit any constraints expected in a highly polyploid bacterium. z

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“…A. vinelandii cells were made competent according to the procedure described previously [22]. In brief, A. vinelandii was cultured on BN Fe 3 agar for about 36 h at 30³C.…”

Section: Preparation Of Competent Cells Of Azotobactermentioning

confidence: 99%

“…This culture was diluted (1:10) in a total volume of 100 ml of BN Fe 3 liquid medium and allowed to grow for 21 h at 30³C by shaking at 200 rpm. Cells prepared in this way were transformed using highly pu-ri¢ed plasmid DNA samples as described previously [22] with the modi¢cations described in Section 3.…”

Section: Preparation Of Competent Cells Of Azotobactermentioning

confidence: 99%

Segregation pattern of kanamycin resistance marker in Azotobacter vinelandii did not show the constraints expected in a polyploid bacterium

Pulakat

1998

FEMS Microbiology Letters

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“…1979) for mobilization. Plate transfonnation of A. vineiandii was based on the method described by Page and von Tigestrom (1979). A. vineiandii cells were grown overnight with vigorous aeration in the transformation medium.…”

Section: Dna Transfermentioning

confidence: 99%

Identification of an operon involved in the assimilatory nitrate‐reducing system of Azotobacter vineiandii

Ramos

Blanco

Gutiérrez

et al. 1993

Molecular Microbiology

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A number of mutants lacking nitrate reductase (Nas-) or nitrite reductase (Nis-) activities have been isolated and characterized. An operon including two new genes (nasA and nasB) has been defined and cloned from an Azotobacter vinelandii gene bank. nasA encodes for nitrite reductase apoenzyme, whereas nasB is specific for nitrate reductase activity. Nitrate reductase exerts a regulatory effect on nasAB.

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“…Azotobacter vinelandii is routinely manipulated using various antibiotic selection markers (Dos Santos 2011; Barney et al 2015). The bacterium is easily made competent and can be transformed with high efficiency (Page and Von Tigerstrom 1979). Additional techniques such as congression offer the opportunity to make manipulations in one region of the genome while adding an antibiotic selection marker to another region of the genome (Robinson et al 1986;Dos Santos 2011).…”

Section: Discussionmentioning

confidence: 99%