A binary vector for transferring genomic libraries to plants.
The transformation of mutant plants with a complete recombinant library derived from wild-type DNA followed by assay of transformed plants for complementation of the mutant phenotype is a promising method for the isolation of plant genes. The small genome of Arabidopsis thaliana is a good candidate for attempting this so-called shotgun transformation. We present the properties of an A. thaliana genomic library cloned in a binary vector, pC22. This vector, designed to introduce genomic libraries into plants, contains the oriV of the Ri plasmid pRiHR1 by which it replicates perfectly stably in Agrobacterium. Upon transfer of the library from E. coli to A. tumefaciens large differences in transfer efficiencies of individual recombinant clones were observed. There is a direct relation between transfer efficiency and stability of the recombinant clones both in E. coli and A. tumefaciens. The stability is independent of the insert size, but seems to be related to the nature of the insert DNA. The feasibility of shotgun transformation and problems of statistical sampling are discussed.
D003001:Cloning, Molecular / Q000379:methods; D004351:Drug Resistance; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D066298:In Vitro Techniques; D007612:Kanamycin / Q000494:pharmacology; D010944:Plants / Q000235:genetics*; D010957:Plasmids; D012098:Reproduction; D012231:Rhizobium / Q000235:genetics*; D014162:Transfection; D014170:Transformation, Genetic
D007612:Kanamycin