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Development of two new molecular markers specific to cytoplasmic male sterility in tuber mustard (Brassica juncea var. tumida Tsen et Lee)

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Abstract

A novel and stable cytoplasmic male sterility CMS line of tuber mustard has been bred by subsequent backcrosses for 10 years. Two specific markers atpA and orf220 were cloned and partially characterized in our previous study (Zhang et al. 2003). In this study, two new molecular markers, orf256 and orf305/orf324, have been isolated and identified. The orf256 gene size was found to be 825 bp in CMS line and a 1,357 bp in its maintainer line. Sequence analysis indicated that the orf256 gene was an entire coding sequence and downstream of the cox1 gene. Interestingly, the 906 bp fragment, which contains part of the sequence of orf222, nad5 and orf139 genes, was found to be inserted from the 451st bp of 5′-flank of the 1,357 bp fragment. In the same way, the orf324 gene was isolated from CMS line and orf305 gene from its maintainer line. Both of them are entire coding sequences, upstream from nad3 and rps12 gene, and co-transcribed with the nad3 and rps12 genes. In addition, two molecular markers, orf256 and orf324/orf305, have been successfully converted into the SCAR markers. Subsequently, ORF256, ORF324, ORF305 protein and ORF256-M-431 fragment are predicated to contain signal peptide sequences, and ORF220 was predicated to contain signal anchor sequence. RFLP analysis results revealed that all of the molecular markers exhibited polymorphisms. Northern blot analysis indicated that the expression level of these genes in CMS line is higher than that of the maintainer line. In the mass, all of these genes are expressed lower in the leaf than that of floral organs between the CMS line and its maintainer line. The difference in expression pattern of different mitochondrial specific marker genes suggests that the abundance of mitochondrial proteins is differentially regulated in the organ/tissue development in tuber mustard. Results of this study also provide some novel and useful clues to explore the biological function of these specific marker genes in the tuber mustard.

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Abbreviations

CMS:

Cytoplasmic male sterility

PCR:

Polymerase chain reaction

RT-PCR:

Reverse transcription polymerase chain reaction

ORF:

Open reading frame

RNAi:

RNA interference

RFLP:

Restriction fragment length polymorphism

SCAR:

Sequence characterized amplified region

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Acknowledgments

This research was partially supported by the Ningbo Science Foundation of China (2004A410018). The authors gratefully acknowledge Dr. Gergely Gulyas for reading and criticizing this manuscript and Dr. Gen Hattori for his helpful advice.

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Authors and Affiliations

  1. Laboratory of Plant Genetic & Breeding, Tokyo University of Agriculture and Technology, Fuchu, Tokyo, 183-8509, Japan

    Xiao-lin Yu, Qiu-bin Xiao & Yutaka Hirata

  2. Laboratory of Cell & Molecular Biology, Institute of Vegetable Science, Zhejiang University, 268 Kaixuan Road, Hangzhou, Zhejiang, 310029, China

    Xiao-lin Yu, Jia-shu Cao & Zhu-jun Chen

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  1. Xiao-lin Yu
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  2. Qiu-bin Xiao
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  3. Jia-shu Cao
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  4. Zhu-jun Chen
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Correspondence to Xiao-lin Yu or Yutaka Hirata.

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Yu, Xl., Xiao, Qb., Cao, Js. et al. Development of two new molecular markers specific to cytoplasmic male sterility in tuber mustard (Brassica juncea var. tumida Tsen et Lee). Euphytica 166, 367–378 (2009). https://doi.org/10.1007/s10681-008-9817-z

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