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Map-based cloning of a recessive genic male sterility locus in Brassica napus L. and development of its functional marker

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Abstract

We previously mapped one male-sterile gene (Bnms3) from an extensively used recessive genic male sterility line (9012AB) in Brassica napus to a 0.14-cM genomic region. In this study, two highly homologous BAC contigs possibly containing the candidate BnMs3 gene were identified using a map-based cloning strategy. A BnMs3-linked SCAR marker (DM1) capable of differentiating the subgenomes between B. rapa and the B. oleracea aided mapping of BnMs3 on the contig derived from the B. napus chromosome C9. One representative BAC clone was sequenced from each of the two contigs and resulted in a larger number of markers according to the sequence difference between the two clones. To isolate BnMs3, these markers were then analyzed in another two BC1 populations with different genetic backgrounds. This assay allowed for a delimitation of the mutated functional region of BnMs3 to a 9.3-kb DNA fragment. Gene prediction suggested that one complete open reading frame (ORF, ORF2) and partial CDS fragments of ORF1 and ORF3 reside in this fragment. Sequence comparison and genetic transformation eventually indicated that ORF1 (designated as BnaC9.Tic40), an analogue of the Arabidopsis gene AT5G16620 which encodes a translocon of the inner envelope of chloroplasts 40 (Tic40), is the only candidate gene of BnMs3. Furthermore, two distinct mutation types in ORF1 both causing the male-sterile phenotype were individually revealed from 9012A and the temporary maintainer line T45. The molecular mechanism of this male sterility as well as the application of BnMs3-associated functional and cosegregated markers in true breeding programs was also discussed.

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Acknowledgments

The authors wish to thank Prof. Jinling Meng for kindly providing the BAC clones of the JBnB BAC library and genomic DNA of Tapidor as well as eight B. oleracea lines. We also thank Prof. Kede Liu for providing the genomic DNA of 50 B. rapa lines used in this research and Dr. Zhixiong Fan for providing the genomic DNA of the temporary maintainer line 9012B-6DH. This research was supported by Natural Science Foundation of China (30670123, 31170166) and National High-tech R&D Program of China (2011AA10A104).

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Authors and Affiliations

  1. National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, 430070, China

    Ji Li, Dengfeng Hong, Junping He, Lei Ma, Lili Wan, Pingwu Liu & Guangsheng Yang

  2. Institute of Industrial Crops, Henan Academy of Agricultural Science, Zhengzhou, 450002, China

    Junping He

  3. Branch of Cultivar Management, Seed Management Station of Hubei Province, Wuhan, 430070, China

    Lei Ma

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  1. Ji Li
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  2. Dengfeng Hong
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  4. Lei Ma
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  6. Pingwu Liu
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  7. Guangsheng Yang
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Correspondence to Guangsheng Yang.

Additional information

Communicated by H. Becker.

J. Li and D. Hong contributed equally to this paper.

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Li, J., Hong, D., He, J. et al. Map-based cloning of a recessive genic male sterility locus in Brassica napus L. and development of its functional marker. Theor Appl Genet 125, 223–234 (2012). https://doi.org/10.1007/s00122-012-1827-5

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