Fast Mapping of a Chlorophyll B Synthesis-Deficiency Gene in Barley (hordeum Vulgare L.) Via Bulked-Segregant Analysis with Reduced-Representation Sequencing

Bulked-segregant analysis coupled with next-generation sequencing（BSA-seq） has emerged as an efficient tool for genetic mapping of single genes or major quantitative trait loci controlling（agronomic） traits of interest. However, such a mapping-by-sequencing approach usually relies on deep sequencing and advanced statistical methods. Application of BSA-Seq based on construction of reduced-representation libraries and allele frequency analysis permitted anchoring the barley pale-green（pg） gene on chromosome 3 HL. With further marker-assisted validation, pg was mapped to a 3.9 Mb physical-map interval. In the pg mutant a complete deletion of chlorophyllide a oxygenase（HvCAO） gene was identified.Because the product of this gene converts Chl a to Chl b, the pg mutant is deficient in Chl b.An independent Chl b-less mutant line M4437 2 carried a nonsynonymous substitution（F263 L） in the C domain of HvCAO. The study demonstrates an optimized pooling strategy for fast mapping of agronomically important genes using a segregating population.