R-plasmid-mediated chromosomal gene transfer in Agrobacterium tumefaciens

Abstract

Although several techniques are available for transferring the Ti plasmids from one strain of Agrobacterium tumefaciens to another, there are no reproducible methods for analysis of chromosomal markers in this phytopathogen. The R plasmid, R68.45, is known to show chromosomal mobilizing ability in several bacterial genera including the closely related Rhizobia. R68.45 was transferred into the prototrophic A. tumefaciens strain 15955. Ten kanamycin-resistant transconjugant clones were tested for chromosomal mobilizing ability by mating with strain SA10, a rifampinand streptomycin-resistant histidine auxotroph of strain 15955. Of the 10 donor clones, 2 showed high chromosomal mobilizing ability. Between 1,000 and 2,000 His' colony-forming units per ml were obtained, 280 on July 31, 2020 by guest

Characterization of transposon Tn5-facilitated donor strains and development of a chromosomal linkage map for Agrobacterium tumefaciens

“…Our transposon-facilitated chromosomal gene transfer system has two advantages over R68.45 in strain 15955, namely polarity and improved mobilization frequencies (8,13,26). Polarized gene transfer systems are particularly useful because, among recombinants selected for acquisition of a distal marker, coinheritance of proximal markers depends only on recombination events.…”

“…Media and chemnicals. Nutrient agar, AB minimal medium, and AB buffer have been previously described (13). Amino acids and antibiotics, purchased from Sigma Chemical Co., were used in the concentrations described previously (26), except that erythromycin was used at 10 ,ug/ml and nalidixic acid was used at 100 jxg/ml.…”

Mutagenesis by insertion of drug resistance transposon Tn7 into a vibrio species

“…The P-1 incompatibility (Inc P1) group R plasmids are unique in showing a broad conjugal host range among gram-negative bacteria. These plasmids have also been shown to mobilize chromosomal transfer in a number of bacterial species, including Pseudomonas aeruginosa (10), Pseudomonas putida (14), Acinetobacter calcoaceticus (19), Rhodopseudomonas sphaeroides (17), Rhizobium leguminosarum (6), and Agrobacterium tumefaciens (11). The present work shows that although RP4 can be transferred by conjugation to this strain of Vibrio, it is not capable of mobilizing the chromosome at a detectable frequency.…”

Intergeneric complementation by Agrobacterium tumefaciens chromosomal genes and its potential use for linkage mapping

“…Mobilization of the Agrobacterium chromosome can be achieved with the Inc P1 plasmid R68.45 at a reasonably high frequency [4][5][6]20].…”