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内容記述 |
Because soybean is a self-pollinating crop and artificial crossing is labor-intensive, we have been developing a recurrent selection method based on insect-mediated pollination using the male sterility gene ms5 and honeybees (Apis mellifera). The male sterility ms5 gene is advantageous for recurrent selection because of the d2 locus, which controls cotyledon color in mature seeds and can be used as a phenotypic selection marker for the ms5 male sterility. However, occasional self-fertilization occurs because of instability of male sterility. Identification of the gene responsible for ms5 male sterility may resolve the problem. We narrowed down the candidate ms5 locus to a 54-kbp region using fine mapping with simple sequence repeat (SSR) markers. Bulked-DNA analysis using next-generation sequencing revealed a deletion and a nucleotide substitution as a candidate variation in the region. The variations were identified in intron of MutS homolog (GmMSH4), which modulates chromosomal recombination in meiosis. The ms5 transcript contained a novel exon with a premature termination codon. This exon originated from an alternative splice acceptor site caused by the deletion and nucleotide substitution, disrupting gene function. Co-segregation of male sterility with five independent mutations in GmMSH4 was confirmed using progeny of mutant lines. Mutations in GmMSH4 led to biased DNA partitioning during meiosis, resulting in collapsed or enlarged pollen and suggesting that ms5 male sterility is caused by the failure of pollen formation during meiosis due to loss of function of GmMSH4. In some genetic backgrounds, normal transcripts were detected in addition to the aberrant transcript, and the ratio between them varied among materials. Although the mechanism underlying the increased production of the normal transcript has not yet been identified, we speculate that an increase in its abundance restores normal pollen formation, thereby enabling self-fertilization. Future studies will aim to identify the conditions that influence transcript composition and to demonstrate the effectiveness of recurrent selection breeding in soybean using the ms5 male sterility system. |
