Genetics of gamma-irradiation-induced mutations in Arabidopsis thaliana: large chromosomal deletions can be rescued through the fertilization of diploid eggs.

Despite the demonstrated value of chromosomal deletions and deficiencies as tools in plant and animal genome research, in the genetic model plant species Arabidopsis thaliana, such mutations have not been extensively studied. For example, it is not known whether large deletions in different regions of the genome can be tolerated in diploid plants that are heterozygous for such mutations. Similarly the viability or inviability of monosomics has not been examined in detail. To investigate these questions, we have used gamma-irradiated haploid wild-type pollen to pollinate diploid and tetraploid multimarker lines of Arabidopsis. Examination of M1 progenies revealed that chromosome loss mutations and large deletions were induced in the irradiated pollen. Such mutations were eliminated in diploid M1 plants due to dominant lethality but could be rescued in triploid M1 progeny. The use of irradiated pollen and tetraploid marker lines of Arabidopsis is a convenient way of generating deletions and modified chromosomes and provides a genetic tool for deletion mapping and for analysis of chromosomal regions essential for chromosome maintenance.

Fine-scale molecular genetic (RFLP) and physical mapping of a 8.9 cM region on the top arm of Arabidopsis chromosome 5 encompassing the male sterility gene, ms1.

Fine-scale molecular mapping has been conducted using 183 recombinants between the markers lutescens (lu; 17.6 cM) and transparent testa glabra (ttg; 35.5 cM) on the top arm of Arabidopsis thaliana chromosome 5. This region contains a number of genes involved in floral development including Ms1, a gene required for the post-meiotic development of pollen. In homozygous ms1 mutant plants, pollen development is aborted soon after microspore release, regardless of environmental conditions. The ms1 mutation is located at 29.8 +/- 0.8 cM on chromosome 5. Markers have been identified which co-segregate with ms1 and should lie within 39 kb of the gene. The fine-scale map of the lu-ms1-ttg region that has been generated is significantly different from the published integrated map and provides substantially more accurate and higher marker density than the current recombinant inbred map for this region. Using clones derived from four yeast artificial chromosome libraries, a contig has been established between the RFLP markers 4111 and 4556, which encompasses the ms1 gene. This covers a genetic distance of 8.9 cM which corresponds to a physical distance of approximately 1.44 Mb, representing about 1.5-2.0% of the Arabidopsis genome. In this region, 1 cM represents a physical distance of approximately 160 kb.

Isolation of deficiencies in the Arabidopsis genome by gamma-irradiation of pollen.

Chromosomal deficiencies are a useful genetic tool in fine-scale genetic mapping and the integration of physical and visible marker genetic maps. Viable overlapping deficiencies may permit gene cloning by subtractive procedures and provide a means of analyzing the functional importance of different chromosomal regions. A method is described for isolation of deficiencies in the Arabidopsis genome which encompass specific loci and other extended chromosomal regions. The technique employs pollen mutagenized by gamma-irradiation to pollinate marker lines homozygous for recessive mutations. Deficiencies at specific loci were detected by screening for marker phenotypes in the F1. Screening for lethal mutations in the F1/F2 confirmed specific deficiencies and revealed other deficiencies that did not overlap the marker loci. Further evidence for such mutations was provided by distorted F2 segregation of the chromosomal markers linked to putative deficiencies. Maintainable (transmissible) and non-transmissible deficiencies were demonstrated by their pattern of inheritance in subsequent generations.