Abstract
Cytoplasmic male-sterility (CMS) has been exclusively used to produce F1 hybrid seeds of onion (Allium cepa L.). A single nuclear locus, Ms, is known to restore male-fertility of CMS in onions. Unstable male-sterile onions producing a small amount of pollen grains have been identified in a previous study. When such unstable male-sterile onions were crossed with stable male-sterile onions containing CMS-R cytoplasm, male-fertility was completely restored, although genotypes of the Ms locus were homozygous recessive. Inheritance patterns indicated that male-fertility restoration was controlled by a single locus designated as Ms2. A combined approach of bulked segregant analysis and RNA-seq was used to identify candidate genes for the Ms2 locus. High resolution melting markers were developed based on single nucleotide polymorphisms (SNPs) detected by RNA-Seq. Comparative mapping of the Ms2 locus showed that Ms2 was positioned at the end of chromosome 2 with a distance of approximately 70 cM away from the Ms locus. Although 38 contigs containing reliable SNPs were analyzed using recombinants selected from 1344 individuals, no contig showed perfect linkage to Ms2. Interestingly, transcription levels of orf725, a CMS-associated gene in onions, were significantly reduced in male-fertile individuals of segregating populations. However, no significant change in its transcription level was observed in individuals of a segregating population with male-fertility genotypes determined by the Ms locus, suggesting that male-fertility restoration mechanism of Ms2 might be different from that of the Ms locus.
Highlights
Male-sterility is an inability of producing functional pollen grains in plants
To investigate whether such an unstable MS phenotype occurred in other types of cytoplasm, two plants (Plants #4 and #9) showing unstable MS phenotypes were crossed with stable MS onions containing the Cytoplasmic male-sterility (CMS)-T cytoplasm (Supplementary Fig. 1)
Genotypes of six molecular markers showed almost perfect linkage disequilibrium with the Ms locus, confirming that genotypes of both MF and MS plants were all homozygous recessive for the Ms locus (Supplementary Fig. 3). These results indicated that male-fertility of these populations was restored by another novel Rf locus, not the Ms locus
Summary
Male-sterility is an inability of producing functional pollen grains in plants. Any critical defects from initial stages of male gametophyte development to anther dehiscence can result in male-sterility (Wan et al 2019). Gynodioecy attributed by male-sterility is a widespread reproductive strategy in flowering plants. 7% of angiosperm species are known to show gynodioecy (McCauley and Bailey 2009). Depending on genomic positions of causal genes, male-sterility is classified as genic male-sterility (GMS) and cytoplasmic male-sterility (CMS). Genes inducing GMS and CMS are located in nuclear and mitochondrial genomes, respectively (Chen and Liu 2013)
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