Agrobacterium-mediated plant transformation by novel mini-T vectors in conjunction with a high-copy vir region helper plasmid.
A new binary vector system for Agrobacterium-mediated plant transformation was developed. A set of four mini-T vectors comprised of T-DNA border sequences from nopaline-type Ti-plasmid pTiC58 flanking a chimaeric hygromycin-resistance gene for selection of transformants and up to eight unique restriction sites for cloning foreign DNA was constructed on a broad-host replicon containing the oriV of plasmid pSa. In two of the constructs these multiple cloning sites are flanked by a strong promoter to activate transcription of inserted DNA in planta. High-efficiency transformation was prompted by a high-copy, stable virulence helper plasmid pUCD2614, which contains a cloned virulence region of pTiC58 and tandem copies of the par locus of plasmid pTAR. Southern blot hybridization and genetic analyses of the progeny of transformed plants showed that the hygromycin resistance gene was stably inherited.
Zyprian
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Department of Plant Pathology, University of California, Davis 95616.
Kado
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D003001:Cloning, Molecular / Q000379:methods*; D004269:DNA, Bacterial / Q000235:genetics; D004351:Drug Resistance; D005822:Genetic Vectors; D006921:Hygromycin B / Q000494:pharmacology; D010947:Plants, Toxic; D010957:Plasmids; D012231:Rhizobium / Q000235:genetics*; D014026:Nicotiana / Q000187:drug effects / Q000235:genetics; D014170:Transformation, Genetic
D004269:DNA, Bacterial; D006921:Hygromycin B