Identification of Rfd1, a novel restorer-of-fertility locus for cytoplasmic male-sterility caused by DCGMS cytoplasm and development of simple PCR markers linked to the Rfd1 locus in radish (Raphanus sativus L.)

Abstract

Previously, novel cytoplasmic male-sterility (CMS) caused by DCGMS cytoplasm was discovered in radish (Raphanus sativus L.) introduced from Uzbekistan. We performed extensive progeny tests and identified two fertility restorer lines ('R171' and 'R121') for this new CMS. Two F 1 hybrid populations were self-pollinated and backcrossed to produce F 2 and BC populations. Inheritance patterns of male-sterility in segregating populations varied depending on paternal lines. Segregation of malesterility in F 2 populations originating from the cross between MS19 and R121 showed that a single locus was involved in fertility restoration. However, populations originating from the cross between MS15 and R171 showed the involvement of more than one restorer-of-fertility genes. The single fertility restorer locus identified in the cross between MS19 and R121 was designated Rfd1 locus. Bulked segregant analysis was performed using RAPD and AFLP, which identified one marker each. Both RAPD and AFLP markers were converted into simple PCR-based co-dominant markers after their isolated flanking sequences were analyzed. Indels 773-bp and 67-bp in length were identified between two Rfd1 allele-linked flanking sequences of the RAPD and AFLP fragments, respectively, then utilized to develop simple PCR markers. In addition, we prove that the newly identified Rfd1 locus is independent of the Rfo locus, another radish fertility restorer for CMS caused by Ogura cytoplasm.

Mitochondrial Genome Sequencing Reveals orf463a May Induce Male Sterility in NWB Cytoplasm of Radish

“…Additionally, DCGMS mitotypes are predominantly in germplasm introduced from eastern European countries [41]. Most breeding lines bred in Korea were identified as maintainers of DCGMS and multiple Rf genes might exist in radish germplasm from European countries [42]. Both NWB CMS and DCGMS lines appear to have the same origin.…”

Construction of a high-resolution linkage map of Rfd1, a restorer-of-fertility locus for cytoplasmic male sterility conferred by DCGMS cytoplasm in radish (Raphanus sativus L.) using synteny between radish and Arabidopsis genomes

“…Ten F 2 individuals consisting of male-sterile and male-fertile bulks were selected based on both the malefertility phenotype and genotypes of the A137 marker, which has been reported to be linked to the Rfd1 locus at a distance of 5 cM (Kim et al 2010). The male-fertile F 2 plants that were heterozygous for the A137 marker were excluded when making DNA bulks.…”

“…Gels were stained with a staining reagent (SILVER SEQUENCE TM , Promega, Madison, WI) according to the manufacturer's instructions. Sequencing of the AFLP fragment linked to the Rfd1 locus was performed as described by Kim et al (2010).…”

“…A novel locus, Rfd1, that restores fertility to male-sterile DCGMS radish plants and a linked marker, A137, were reported in our previous study (Kim et al 2010). To develop additional linked markers for the construction of a linkage map of the Rfd1 locus, the BSA approach was used in combination with AFLP analysis.…”

“…Fertility restoration was conditioned by a single locus, Rfd1, and a linked molecular marker was developed (Kim et al 2010). However, construction of a high-resolution linkage map was not developed due to a lack of radish genomic information.…”