Two complementary restorer-of-fertility loci confer the pollen fertility restoration in the progenies of ‘Niitaka’ pear (Pyrus pyrifolia)

• Morphological and genetic characterization of cytoplasmic male sterility in ‘Niitaka’-derived pears. • Two-locus complementary model explaining the 3:5 segregation ratio. • Fine mapping of Rf1 (Chr4) and Rf2 (Chr5) using SNP and InDel markers. • Identification of PPR candidate genes linked to fertility restoration. Cytoplasmic male sterility (CMS) refers to the failure of flowering plants to produce viable pollen grains during anther development. CMS and the restorer-of-fertility ( Rf ) system have been documented in Pyrus . Male sterility has been reported in the Asian pear ‘Niitaka’ ( P. pyrifolia ) and its F 1 progeny. However, Rf genes conferring pollen fertility for ‘Niitaka’ cytoplasm in pears remains to be elucidated. Therefore, this study aimed at precisely mapping Rf genes involved in pollen fertility restoration and exploring candidate genes. We analyzed F 1 progenies derived from female ‘'Whangkeumbae' ( P. pyrifolia ) possessing ‘Niitaka’ cytoplasm. Two non-linked loci with complementary dominance were hypothesized to account for the observed separation ratio. Genotyping-by-sequencing (GBS) analysis was performed to identify single nucleotide polymorphism (SNP) markers and finely map the Rf loci. Two Rf loci were preliminarily mapped to linkage groups 4 and 5 and named Rf1 and Rf2 , respectively. Insertion/deletion (InDel) markers were developed by analyzing the whole genome sequencing data of the parents. Rf1 and Rf2 regions were delimited to 560 kb and 880 kb, respectively, by fine mapping with InDel markers. Four candidate genes encoding pentatricopeptide repeat proteins were identified in Rf1 and Rf2 . To the best of our knowledge, this is the first study to map Rf genes responsible for cytoplasmic male sterility in pears, which advances our understanding of male sterility.