ABSTRACT
Plant Stress & Germplasm Development Unit, Cropping Systems Research Laboratory, USDAARS, 3810 4th St., Lubbock, TX, USA, 79415. Email: John.burke@ars.usda.gov *Corresponding author: Zhanguo.xin@ars.usda.gov
Many important signal transduction components, developmental regulators and biochemical pathways are discovered through forward genetic analysis of mutants in model plants (https://thearabidopsisbook.org). The process begins with mutagenesis of a pure (inbred) line with either a physical or chemical mutagen and isolation of mutants with relevant phenotypes. The mutant of interest is outcrossed to another accession that has extensive DNA polymorphism to the line used for mutagenesis. Linkage of the mutant phenotype with DNA markers is analyzed using as many markers as possible until the mutation is mapped into a very small region fl anked by two DNA markers. The process, called map-based cloning, depends on two critical factors to succeed: a large segregating mapping population and dense DNA markers to have the resolution power to delimit the mutation into a small region. Usually, over 1,000 individual F2 plants with an accurate phenotype need to be analyzed with hundreds of DNA markers in order to narrow the mutation to a region harboring only a few genes (Jander et al. 2002). The entire region is sequenced to identify the gene that carries the expected mutation. To confi rm the identity of the gene, the wild type gene is introduced into the mutant through transformation to determine if the gene can restore the mutant to wild type phenotype. This last step can be by-passed if two or more independent mutant alleles are identifi ed that carry unique mutations in the same gene.
