Molecular genetic analysis of pollen irradiation mutagenesis in Arabidopsis

Abstract

• Pollen γ-irradiation mutagenesis was systematically investigated to generate targeted mutations in Arabidopsis. • Irradiation effects on viability, germination and frequency of loci deletions were evaluated. • Mutation frequency increased with irradiation dose and varied depending upon pollen developmental stage. Meiosis was the most irradiation-sensitive stage, however, it did not equate to the highest mutation frequency. High frequencies of targeted mutations were obtained by irradiation from the second mitosis to mature pollen stages, using 400-600 Gy. Targeted mutations could also be obtained using lower doses of γ-rays (e.g. 200 Gy) provided that pollen was irradiated at an earlier developmental stage. ms1ttg marker locus pseudo-dominants were used to verify the presence and size of the resultant deletions. • The results demonstrate that γ-irradiation of pollen is an efficient approach for generating deletions in the Arabidopsis genome. Pollen mutagenesis offers the possibility of combining single-cell selection procedures with the advantages of haploid systems, including the ability to treat large numbers of pollen grains, the absence of chimerism and direct expression of targeted alleles in the M1 generation. © New Phytologist (2004).