We constructed a cosmid vector, pOCA18, designed for transferring plant genomic libraries from A grobacterium tumefaciens to plants. Clones from a genomic library of Arabidopsis thaliana DNA in pOCA18 were propagated stably in both Escherichia coli and A. tumefaciens. Clones from the pOCA18 A. thaliana library were used to construct transgenic Nicotiana tabacum plants; the DNA inserts were transferred intact in 10 out of 16 transgenic N. tabacum plants examined but were partially deleted in six others. Transgenic N. tabacum plants constructed with a mutant A. thaliana acetohydroxy acid synthase gene (from the pOCA18 library) that encodes an enzyme resistant to the herbicide chlorsulfuron were resistant to chlorsulfuron. A statistical analysis indicated that if the A. thaliana library contains 107 members and if 107 A. tumefaciens transconjugants containing the library were used to transform plant cells, then 2 × 104 transformed plant cells must be generated to have a 95% probability of constructing a transgenic plant carrying a specific DNA sequence from the A. thaliana library..Specialized binary vector for plant transformation: expression of the Arabidopsis thaliana AHAS gene in Nicotiana tabacum
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ACKNOWLEDGMENTS We thank B. Lalonde and G. Fink for providing pBL104 containing the wild-type A. thaliana AHAS gene prior to publication, G. Haughn and C. Somerville for providing A. thaliana line GH-50, M. Stempowski and J. Smith for synthesizing the left border oligonucleotide, R. Hyde for assistance in the preparation of this manuscript, and J. Chory for helpful discussions and donating the C to pOCA. This work was supported by a grant from Hoechst AG to Massachusetts General Hospital.
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