Somatic cell selection for chlorsulfuron-resistant mutants in potato: identification of point mutations in the acetohydroxyacid synthase gene

Abstract Background Somatic cell selection in plants allows the recovery of spontaneous mutants from cell cultures. When coupled with the regeneration of plants it allows an effective approach for the recovery of novel traits in plants. This study undertook somatic cell selection in the potato (Solanum tuberosum L.) cultivar ‘Iwa’ using the sulfonylurea herbicide, chlorsulfuron, as a positive selection agent. Results Following 5 days’ exposure of potato cell suspension cultures to 20 μg/l chlorsulfuron, rescue selection recovered rare potato cell colonies at a frequency of approximately one event in 2.7 × 105 of plated cells. Plants that were regenerated from these cell colonies retained resistance to chlorsulfuron and two variants were confirmed to have different independent point mutations in the acetohydroxyacid synthase (AHAS) gene. One point mutation involved a transition of cytosine for thymine, which substituted the equivalent of Pro-197 to Ser-197 in the AHAS enzyme. The second point mutation involved a transversion of thymine to adenine, changing the equivalent of Trp-574 to Arg-574. The two independent point mutations recovered were assembled into a chimeric gene and binary vector for Agrobacterium-mediated transformation of wild-type ‘Iwa’ potato. This confirmed that the mutations in the AHAS gene conferred chlorsulfuron resistance in the resulting transgenic plants. Conclusions Somatic cell selection in potato using the sulfonylurea herbicide, chlorsulfuron, recovered resistant variants attributed to mutational events in the AHAS gene. The mutant AHAS genes recovered are therefore good candidates as selectable marker genes for intragenic transformation of potato

Genomic data from the potato

Available here is the genome of the potato (Solanum tuberosum L.), the first genome sequenced from the asterid clade. Potato is a member of the Solanaceae, a plant family that includes many other economically important species, such as tomato, petunia, eggplant, tobacco, and pepper. As the potato is both clonally propagated and the world;s most important non-grain food crop, its genome is a valuable agricultural resource. The Potato Genome Sequencing Consortium sequenced two species: the heterozygous diploid S. tuberosum Group Tuberosum cultivar, RH89-039-16 (RH), and the doubled monoploid S. tuberosum Group Phureja clone DM1-3 516R44 (DM). The potato genome consists of 12 chromosomes, of which over 80% of the homozygous clone’s 844-megabase genome were assembled. Genome analysis revealed evidence of at least two genome duplication events and identified a number of asterid-specific genes. Comparison between the two clones identified frequent gene variations and mutations, which may cause inbreeding depression