Development of a molecular marker specific to a novel CMS line in radish (Raphanus sativus L.).
ABSTRACT In this study, we have investigated the cytoplasmic male sterility (CMS) of a novel male sterile radish line, designated NWB CMS. The NWB CMS was crossed with 16 fertile breeding lines, and all the progenies were completely male sterile. The degree of male sterility exhibited by NWB CMS is more than Ogura CMS from the Cruciferae family. The NWB CMS was found to induce 100% male sterility when crossed with all the tested breeding lines, whereas the Ogura CMS did not induce male sterility with any of the breeding lines. PCR analysis revealed that the molecular factor that influenced Ogura CMS, the orf138 gene, was absent in the NWB CMS line, and that the orf138 gene was not also expressed in this CMS line. In order to identify the cytoplasmic factors that confer male sterility in the NWB CMS line, we carried out RFLP analyses with 32 mitochondrial genes, all of which were used as probes. Fourteen genes exhibited polymorphisms between the NWB CMS line and other radish cultivars. Based on these RFLP data, intergenic primers were developed in order to amplify the intergenic regions between the polymorphic genes. Among these, a primer pair at the 3' region of the atp6 gene (5'-cgcttggactatgctatgtatga-3') and the 5' region of the nad3 gene (5'-tcatagagaaatccaatcgtcaa-3') produced a 2 kbp DNA fragment as a result of PCR. This DNA fragment was found to be specific to NWB CMS and was not present in other CMS types. It appears that this fragment could be used as a DNA marker to select NWB CMS line in a radish-breeding program.
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ABSTRACT: We analysed DNA and complementary DNA (cDNA) from two inbred lines (CMS-ARIT121 and CMS-ARIT2) along with their respective maintainer lines with eight known genes (atpa, atp6, atp9, coxI, coxII, orfB, orf222 and orf224) reported in other cytoplasmic male sterility (CMS) systems. DNA analyses indicated that only orf224 was different in the maintainer and CMS in ARIT121. Sequencing analysis indicated that CMS-ARIT121 has a similar sequence to that of polima CMS. However, CMS-ARIT2 was quite unique, with no sequence difference at DNA or cDNA level in all eight candidate genes, with the exception of a two-base substitution in the atp6 gene between the CMS and maintainer lines. The similar amplification pattern with all primers, especially orf224, indicates that CMS-ARIT2 is unique and does not correspond to polima, ogura or nap. However, further research is required to establish the causes of sterility in this cms, since we could not find any significant difference at transcript level between CMS-ARIT2 and its corresponding maintainer line. However, the same restorer PR121 as for CMS-ARIT121 was able to restore CMS-ARIT2, and as such can be used for hybrid seed production in Brassica napus L. Overall, this study provides important information for breeders selecting new cytoplasmic male sterility (CMS) in B. napus L. and use of molecular markers to identify CMS lines at early seedling stage that will enhance the B. napus L. breeding program.Euphytica 12/2014; 200(3). DOI:10.1007/s10681-014-1195-0 · 1.69 Impact Factor
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ABSTRACT: Brassicaceae crops display strong hybrid vigor, and have long been subject to F1 hybrid breeding. Because the most reliable system of F1 seed production is based on cytoplasmic male sterility (CMS), various types of CMS have been developed and adopted in practice to breed Brassicaceae oil seed and vegetable crops. CMS is a maternally inherited trait encoded in the mitochondrial genome, and the male sterile phenotype arises as a result of interaction of a mitochondrial CMS gene and a nuclear fertility restoring (Rf) gene. Therefore, CMS has been intensively investigated for gaining basic insights into molecular aspects of nuclear-mitochondrial genome interactions and for practical applications in plant breeding. Several CMS genes have been identified by molecular genetic studies, including Ogura CMS from Japanese radish, which is the most extensively studied and most widely used. In this review, we discuss Ogura CMS, and other CMS systems, and the causal mitochondrial genes for CMS. Studies on nuclear Rf genes and the cytoplasmic effects of alien cytoplasm on general crop performance are also reviewed. Finally, some of the unresolved questions about CMS are highlighted.Breeding Science 05/2014; 64(1):38-47. DOI:10.1270/jsbbs.64.38 · 1.34 Impact Factor
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ABSTRACT: Protoplasts from cabbage and radish were isolated and fused symmetrically by PEG treatment. The PEG treated mixture of high concentrated protoplasts produced lots of micro-calli after weeks. The microcalli developed to normal calli and shoots were regenerated from the calli. A total of 218 shoots were regenerated, but none of them contained the NWB-CMS specific DNA marker, indicating that the transfer of the radish NWB-CMS character into cabbage did not occur. However, ISSR analysis revealed that the cell fusion between protoplasts from radish and cabbage was occurred (3 out of 208 plantlet). The fused regenerants possessed the characteristics of source plants used for protoplast fusion. After vernalization, three regenerants were flowered with white petal color as seen in radish. Only three seeds were able to obtain from one regenerant by backcrossing with the cabbage pollen.01/2008; 35(2). DOI:10.5010/JPB.2008.35.2.121