Identification of the predominant nonrestoring allele for Owen-type cytoplasmic male sterility in sugar beet (Beta vulgaris L.): development of molecular markers for the maintainer genotype

Abstract

Hybrid seed production in sugar beet relies on cytoplasmic male sterility (CMS). As time-consuming and laborious test crosses with a CMS tester are necessary to identify maintainer lines, development of a marker-assisted selection method for the rf gene (the nonrestoring allele of restorer-of-fertility locus) is highly desirable for sugar-beet breeding. To develop such a method, we investigated genetic variation at the Rf1 locus, one of two Rf loci known in sugar beet. After HindIII-digestion, genomic DNAs from beet plants known to have a restoring Rf1 allele yielded a range of hybridization patterns on agarose gels, indicating that Rf1 is a multi-allelic locus. However, the hybridization patterns of 22 of 23 maintainer lines were indistinguishable. The nucleotide sequences of the rf1 coding regions of these 22 maintainer lines were found to be identical, confirming that the lines had the same rf1 allele. Two PCR markers were developed that targeted a downstream intergenic sequence and an intron of Rf1. The electrophoretic patterns of both markers indicated multiple Rf1 alleles, one of which, named the dd(L) type, was associated with the maintainer genotype. To test the validity of marker-assisted selection, 147 sugar beet plants were genotyped using these markers. Additionally, the 147 sugar beet plants were crossed with CMS plants to determine whether they possessed the maintainer genotype. Analysis of 5038 F1 offspring showed that 53 % of the dd(L) plants, but none of the plants with other alleles, had the maintainer genotype. Thus, selection for the dd(L) type considerably enriched the proportion of plants with the maintainer genotype.Electronic supplementary materialThe online version of this article (doi:10.1007/s11032-013-9854-8) contains supplementary material, which is available to authorized users.

Post‐translational mechanisms are associated with fertility restoration of cytoplasmic male sterility in sugar beet (Beta vulgaris)

“…Our analysis included another transgene made from bvORF20L , the bvORF20 counterpart of rf1rf1 sugar beet lines (Matsuhira et al ., ; Moritani et al ., ). Although levels of transcripts and the translation products of bvORF20L were below the limits of detection by RNA gel blot analysis or immunoblot analysis (Matsuhira et al ., ; and this study), transcription of bvORF20L was indicated by RT‐PCR analysis (Fig.…”

Molecular Cloning of X Locus and Marker-Assisted Selection of Non-Restoring Allele for Owen Cytoplasmic Male Sterility

“…However, 22 of 23 maintainer lines showed the same hybridization pattern. The sequences of the rf1 coding regions of these 22 maintainer lines were found to be identical, confirming that they shared the same rf1 allele (Moritani et al, 2013). Two PCR markers targeting a downstream intergenic sequence and the first intron of Rf1, respectively, were developed.…”

“…The PCR protocol was 35 cycles of 94˚C for 3 s , 58˚C for 1 min and 72˚C for 1 min. The DNA fragment used for preparation of the DNA probe was PCR amplified using the primers shown in Moritani et al (2013).…”

The use of genetic, manual and chemical methods to control pollination in vegetable hybrid seed production: a review

“…The most commonly used cytoplasm is Owen type originally described in sugar beet as the result of the interaction between a sterilizing cytoplasm and at least two nuclear restorer genes and environmental factors (Owen 1942(Owen , 1945. Restorer gene Rf1 has been cloned and sequenced (Hagihara et al 2005, Matsuhira et al 2012, and molecular markers are available for MAS (Moritani et al 2013). Recently, Honma et al (2014) have reported the molecular mapping of Rf2.…”