Agrobacterium tumefaciensT-DNA Integration and Gene Targeting inArabidopsis thalianaNon-Homologous End-Joining Mutants

[摘要] In order to study the role of AtKu70 and AtKu80 inAgrobacterium-mediated transformation and gene targeting, plant lines with a T-DNA insertion inAtKu80orAtKu70genes were functionally characterized. Such plant lines lacked both subunits, indicating that heterodimer formation between AtKu70 and AtKu80 is needed for the stability of the proteins. Homozygous mutants were phenotypically indistinguishable from wild-type plants and were fertile. However, they were hypersensitive to the genotoxic agent bleomycin, resulting in more DSBs as quantified in comet assays. They had lower end-joining efficiency, suggesting that NHEJ is a critical pathway for DSB repair in plants. BothAtkumutants and a previously isolatedAtmre11mutant were impaired inAgrobacteriumT-DNA integration via floral dip transformation, indicating that AtKu70, AtKu80, and AtMre11 play an important role in T-DNA integration inArabidopsis. The frequency of gene targeting was not significantly increased in theAtku80andAtku70mutants, but it was increased at least 10-fold in theAtmre11mutant compared with the wild type.

[学科分类] 植物学