Arabidopsis 2010: Saturating the Arabidopsis Knockout Collection with Lethals

Oklahoma State University, Stillwater OK

Investigators

Abstract

One goal of the Arabidopsis research community is to identify a knockout for every annotated gene. Missing from the current collection of homozygous knockout mutants are the essential genes for which mutant seeds abort or fail to form because of gametophytic lethality. The aim of this project is to coordinate efforts to fill this gap in the knockout collection. The result will be a valuable resource for future experiments and benchmark information on the consequences of disrupting non-redundant, essential genes in Arabidopsis. The principal investigators have already established a robust database (www.seedgenes.org) of 300 EMB genes required for seed development in Arabidopsis. Three approaches are planned to expand upon this community resource. The first involves genes where the existing knock-out project finds insertion lines with confirmed heterozygotes but no homozygous knockouts. This list will be filtered to identify genes that represent promising candidates for embryo or gametophytic lethals based on known sequence redundancy, orthology, expression patterns, and shared features with known essential genes. Selected lines will then be planted, genotyped, and confirmed. They will be screened for defects in seed and gametophyte development. The goal is to analyze 450 to 600 promising insertion lines over three years. The second approach involves collecting information on lethal mutants identified in other laboratories. This approach has worked well for seed mutants and will expand in the future to include putative gametophytic mutants. The third approach involves identification of promising candidates for essential genes among those remaining genes for which insertions have not been found or cannot be confirmed. This approach will generate a list of candidate genes that are required for both male and female gametophyte development. Results of these combined approaches will be incorporated into the SeedGenes and TAIR databases. Genes with unknown functions and without orthologs in other model organisms but with a confirmed seed phenotype will be examined in most detail. These efforts will enhance our understanding of the unknowns in Arabidopsis by focusing on genes known to be essential. The broader educational impact of this project includes participation of high school students in research activities and outreach programs on Arabidopsis mutants and plant molecular genetics involving regional high schools, the Extended Studies Program at Stillwater Junior High School, and the Oklahoma School for Science and Mathematics in conjunction with the OSU Plant Biotechnology Network. These programs were successfully initiated during the previous grant period.