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An Arabidopsis histone H2A mutant is deficient in Agrobacterium T-DNA integration

Mysore, K.S., Nam, J., Gelvin, S.B.
Proceedings of the National Academy of Sciences of the United States of America 2000 v.97 no.2 pp. 948-953
Arabidopsis thaliana, transgenic plants, genetic transformation, roots, Agrobacterium tumefaciens, histones, mutants, gene expression, reporter genes, nucleotide sequences, genes, genetic complementation, genetic recombination, transfer DNA
Agrobacterium tumefaciens genetically transforms plant cells by transferring a portion of the bacterial Ti-plasmid, the T-DNA, to the plant and integrating the T-DNA into the plant genome. Little is known about the T-DNA integration process, and no plant genes involved in integration have yet been identified. We characterized an Arabidopsis mutant generated by T-DNA insertional mutagenesis, rat5, that is resistant to Agrobacterium root transformation. rat5 contains two copies of T-DNA integrated as a tandem direct repeat into the 3' untranslated region of a histone H2A gene, upstream of the polyadenylation signal sequence. Transient and stable beta-glucuronidase expression data and assessment of the amount of T-DNA integrated into the genomes of wild-type and rat5 Arabidopsis plants indicated that the rat5 mutant is deficient in T-DNA integration. We complemented the rat5 mutation by expressing the RAT5 histone H2A gene in the mutant plant. Overexpression of RAT5 in wild-type plants increased Agrobacterium transformation efficiency. Furthermore, transient expression of a RAT5 gene from the incoming T-DNA was sufficient to complement the rat5 mutant and to increase the transformation efficiency of wild-type Arabidopsis plants.