High-throughput Genotyping Of Rice Recombination Populations By Next-generation Sequencing And Chromosome 4 Comparison Of Oryza Sativa Ssp. Indica And Japonica Genomes

Cultivated rice is one of the principle food supplies of the world. Nearly half of the world population's staple food is rice. The Asian cultivated rice (Oryza sativa) has two major subspecies: indica and japonica. Sequencing and comparative analysis of indica and japonica genomes are essential to understand their genetic variation. We sequenced ~22.1 Mb regions on chromosome 4 of indica Guangluai-4 for comparing with the corresponding regions on chromosome 4 of japonica Nipponbare. We identified 81,050 single nucleotide polymorphisms, 17,550 insertions or deletions, and 27,889 non-homologous segments. We also analyzed the repeat sequences, alternative slicing events in genes, and subspecies specific genes. These findings give important insights into natural selection and domestication of rice, and provide important resources for rice breeding, molecular cloning, and evolutionary studies. Genetic mapping using recombination populations of indica and japonica offer a way to relate genetic variation to their phenotypic variation between the two subspecies. The conventional techniques greatly restricted mapping resolution for quantitative trait locus (QTL) in recombination populations. The advent of the second-generation sequencing technique has set the stage for replacing the PCR-based approach with the sequencing-based strategy. We developed a high-throughput method for genotyping recombinant populations utilizing whole-genome resequencing data generated by the Illumina Genome Analyzer (GA). A sliding window approach is designed to collectively examine genome-wide single nucleotide polymorphisms for genotype calling and recombination breakpoint determination. This sequencing-based high-throughput genotyping method combines advantages of time and cost effectiveness, high mapping accuracy and resolution. To facilitate a wide application of this new method, we further developed a pipeline, SEG-Map (Sequencing Enabled Genotyping for Mapping recombination populations), that allows the direct analysis of Illumina GA short reads for map construction. By using SEG-Map software package, we perform the sequencing-based genotyping in the populations of rice recombinant inbred lines (RILs), rice chromosome segment substitution lines (CSSLs), rice F2 population. High-resolution and high-accuracy recombination maps were able to be constructed in all the populations, with QTL mapped in a relatively high resolution.