Novel constructions to enable the integration of genes into the Agrobacterium tumefaciens C58 chromosome

Molecular Plant-microbe Interactions : MPMI
Lan-Ying LeeS B Gelvin

Abstract

We constructed several versatile sets of vectors that can be used to introduce any gene into the pgl/picA locus of the Agrobacterium tumefaciens C58 chromosome without affecting T-DNA transfer. One set contains a fragment containing the lacIq and lacZ genes and a multiple cloning site from pBluescriptII SK(+) inserted into a PstI site between the pgl and picA genes on an incPalpha plasmid. The resulting plasmid contains eight unique restriction endonuclease sites and the ability to use blue-white screening for the presence of an insert. A second plasmid also contains a beta-lactamase gene within this locus and provides a convenient ampicillin-carbenicillin resistance marker for the selection of genes integrated into the chromosome following double homologous recombination (homogenotization). A third plasmid contains, in addition to the lacZ, lacIq, and beta-lactamase genes within the pgl/picA locus, a sacRB gene cassette within the vector to counterselect against the presence of the vector within A. tumefaciens. To test this system, we introduced a wild-type virD2 gene into the A. tumefaciens chromosome at the pgl/picA locus. When a Ti plasmid harboring a deletion of virD2 was in this strain, the integrated virD2 gene complemen...Continue Reading

Citations

Feb 6, 2008·Plant Physiology·Lan-Ying Lee, Stanton B Gelvin
Mar 11, 2003·Microbiology and Molecular Biology Reviews : MMBR·Stanton B Gelvin
Aug 10, 2021·The Plant Journal : for Cell and Molecular Biology·Philippe V JutrasRenier A L van der Hoorn

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