Effects on Mapping the Gene of Green-revertible Chlorina 2 (grc2) Using SLAF-seq BSA in Rice (Oryza sativa)
TAN Yan-Ning1, YU Dong1, 2, SHENG Xia-Bing1, 2, KANG Wei-Wei2, LI Zhe-Li1, 3, DUAN Mei-Juan3, YUAN Ding-Yang1, 3, *
1 State Key Laboratory of Hybrid Rice, Hunan Hybrid Rice Research Center, Changsha 410125, China; 2 College of Biosciences and Biotechnology, Hunan Agricultural University, Changsha 410128, China; 3 College of Agronomy, Hunan Agricultural University, Changsha 410128, China
Abstract：SLAF-seq BSA, integrated traditional bulk segregant analysis (BSA) with specific-locus amplified fragment sequencing (SLAF-seq), makes it possible to quickly locate qualitative trait genes. However, little is known about the reliability of its mapping results. In this study, the green-revertible chlorina gene (grc2) in rice (Oryza sativa) was selected as a target for comparing the mapping results provided with map-based mapping and SLAF-seq BSA. Firstly, 5302 SLAF markers were identified which showed single nucleotide polymorphism between 'grc2' and 'Y58S' (a cultivar presenting normally green leaves) by SLAF-seq. Later, the wild-type bulk and the mutant bulk were established, each consisting of 100 plants from a F2 population of 'Y58S'/'grc2'. By automatically calculating the genotype frequency in two pools, 13 SLAF markers were found associated with grc2 which accorded with an expected separation ratio of 33∶66 for paternal / maternal in wild-type bulk and 99∶1 in mutant bulk at 0.01 level. Interestingly, all of the associated markers were distributed on two neighbour regions of 0.98~1.21 Mb and 1.57~1.99 Mb on Chromosome 6 referenced as '93-11' (an indica cultivar). Furthermore, two markers named AM1 and AM6 on the first region were confirmed linkaged with grc2, just exhibiting a genetic distance of 0.625 cM and 0 cM respectively. While referenced as 'Nipponbare' (a japonica cultivar), it was observed that the first region just covered the mapping area with traditional method, indicating the result of SLAF-seq BSA is for consulting. Therefore, the results presented herein supported SLAF-seq BSA would be a useful tool for rapidly obtaining the region associated with qualitative trait gene.
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