Scientia Agricultura Sinica ›› 2015, Vol. 48 ›› Issue (4): 621-629.doi: 10.3864/j.issn.0578-1752.2015.04.01

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS •     Next Articles

Genetic Analysis and Mapping of the Rice Male Sterile Mutant oss125

ZHANG Wen-hui1, YAN Wei2, CHEN Zhu-feng1, XIE Gang1, LU Jia-wei1, LIU Dong-feng1, TANG Xiao-yan1,2   

  1. 1Shenzhen Institute of Molecular Crop Design, Shenzhen 518107
    2College of Life Sciences, Capital Normal University, Beijing 100089
  • Received:2014-08-08 Online:2015-02-16 Published:2015-02-16

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Abstract: 【Objective】This study is to conduct genetic analysis on the rice male sterility mutant oss125 and clone the mutant gene using modified MutMap method, laying a foundation for further study of the gene function and its application in agricultural production. 【Method】 A male sterility mutant, designated as oss125, was isolated from a mutant library of the indica rice Huanghuazhan (HHZ) generated with ethyl methane sulfonate (EMS) treatment. Phenotype of the oss125 mutant was observed. The oss125 mutant was crossed with the wild-type HHZ, fertility of the F1 DNA of 30 F2plants showing the male sterile phenotype were isolated and bulked in equal amount. Bulked DNA was subjected to whole-genome re-sequencing on Hiseq 2000 platform. A modified MutMap method was utilized to identify genomic regions harboring the causal mutation. Linkage of candidate genes with the mutant phenotype was validated with HRM analysis. Sequence analysis of the candidate gene and the expression of the candidate gene in various rice tissues were studied using real-time PCR. 【Result】 Compared with the wild type HHZ, oss125 mutant had normal vegetative growth, inflorescence and flower morphology. 1% I2-KI staining revealed 85% abnormal pollen and 15% pollen of normal phenotype. But the plant displayed complete male sterility. All F1 plants derived from the cross of oss125 mutant with the wild-type HHZ were fertile, and the F2 plants displayed 3:1 segregation of fertile to male sterile plants, indicating that oss125 is controlled by a single recessive gene. Using a modified MutMap method, four candidate SNPs were identified, three in intergenic region, and one in the second exon of OsRPA1a. A single nucleotide mutation (A663C) occured in the coding region of the OsRPA1a gene, resulting in a nonsynonymous mutation (Q221P) in protein sequence. Linkage of the mutation with the male sterile phenotype was validated using HRM analysis. 【Conclusion】 Mutation of the OsRPA1a gene is responsible for the oss125 mutant phenotype. A663C mutation causes abnormal pollen development, leading to complete male sterility. OsRPA1a is essential for meiosis and DNA repair in somatic cells. It was previously reported that a T-DNA insertion mutant of OsRPA1a was reduced in male fertility but completely female sterile. However, oss125 mutant is completely male sterile, and the female fertility appears not affected, suggesting that OsRPA1a involves distinct regions in controlling male and female gametophyte development. The mutation in the oss125 mutant may be in the region governing male gametophyte development but not affecting the function of the female gametophyte development.and F2 plants was investigated.

Key words: rice, mutant, male sterility, HRM, OsRPA1a

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