Isolation of Rice Genes Involved in Important Agronomic Traits Using Whole Genome Resequencing

Over 10,000 mutant lines (M3 to M5) were developed by EMS treatment of two japonica rice cultivars, Hitomebore and Sasanisiki. These mutant lines are invaluable research material to isolate genes and ultimately decipher gene functions. Many interesting agro-morphological traits are depicted by these mutant lines, which have already been used for the isolation of few interesting genes. In this series, two mutant lines namely Hit5369 and Hit1227 were selected that showed early senescence and dwarf and narrow leaf phenotypes, respectively. Crosses were made between these identified mutants and an indica rice cultivar Kasalath. Segregation of the F2 progeny were scored in the paddy field and F2 plants showing the mutant phenotypes were then employed for rough-mapping using SSR markers. For fine mapping, new InDel markers were developed for the regions where polymorphic SSR markers were not available. For Hit5369 (early senescence), the candidate locus was narrowed down to 365 kb region on chromosome 1, which is predicted to contain 52 genes. Illumina whole genome re-sequencing of the mutant is currently underway to identify the causative mutation by comparing the mutant sequence to Hitomebore reference sequence within the candidate region. For Hit1227 (dwarf and narrow leaf), the mutation is mapped to about 700Kb on chromosome 4, a region that also harbors an already cloned geneNal1.  Sequencing of this gene in Hit1227 and wild-type Hitomebore revealed a SNP in the coding region that caused amino acid change from Thr (ACC) to Ile (ATC), indicating that Hit1227 is an allele of the Nal1.