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Euphytica

, Volume 181, Issue 1, pp 17–29 | Cite as

Molecular characterization of cytoplasmic male sterility conditioned by Gossypium harknessii cytoplasm (CMS-D2) in upland cotton

  • Jianyong Wu
  • Yangcang Gong
  • Minghui Cui
  • Tingxiang Qi
  • Liping Guo
  • Jinfa Zhang
  • Chaozhu XingEmail author
Article

Abstract

Cytoplasmic male sterility (CMS) is a maternally inherited trait that fails to produce functional pollen grains. The CMS system is widely employed to facilitate the utilization of heterosis in major crops. However, little is known about the CMS associated genes in Upland cotton (Gossypium hirsutum). The objective of this study was to compare CMS cotton (CMS-D2) with the cytoplasm from G. harknessii and its isogenic maintainer line with the normal fertile Upland cotton cytoplasm to identify CMS-D2 specific gene(s) and to develop CMS-specific sequence characterized amplified region (SCAR) markers. Based on Southern blot analysis using 10 mitochondrial gene-specific probes (cob, cox2, atp6, atp9, nad3, cox3, atpA, cox1, nad6 and nad9), three probes (cox3, atpA, and nad6) revealed restriction fragment length polymorphisms (RFLP) between the CMS-D2 and its isogenic maintainer line. RT-PCR confirmed that the three genes were differentially expressed between the two lines. These results indicated that there existed structural and expression variations in the three genes when the mitochondrial D2 genome was transferred into Upland cotton. Genome walking and rapid amplification of cDNA ends (RACE) were further performed to analyze the sequences of these genes and their flanking regions. For cox3 and nad6, there was only one different nucleotide each in the gene regions between the two lines. Also some nucleotides upstream of the ATG codon were different. For atpA, the sequences downstream the atpA were significantly different between the two cytoplasmic lines. Furthermore, two nucleotides at the -4 and -5 position from ATG codon were also changed between the two cytoplasms (i.e., CG→AA), and this mutation also exists in RNA sequences. Interestingly, nine nucleotides (ATGCAACTA) were also inserted at the location of 899 bp upstream of ATG codon in the CMS line. The results suggest that the abnormal sequence and expression of atpA gene is associated with CMS expression in Upland cotton. According to the significant different sequences downstream the atpA gene, a CMS-D2 specific SCAR marker was developed. The CMS-specific PCR bands were verified for 10 cultivars containing either normal- or CMS-D2cytoplasm. This will allow quick and reliable identification of the cytoplasmic types of individual plants at the seedling stage, and assessment of the purity of F1 seed lots.

Keywords

Upland cotton CMS atpA ATG codon SCAR 

Notes

Acknowledgment

This research was financed by funds from the National Transgenic Major Program (2008ZX08005-001) and Special Fund for Basic Research in the Central Public Research Institutes (SJA0903).

Supplementary material

10681_2011_357_MOESM1_ESM.doc (4.4 mb)
Supplementary material 1 (DOC 4508 kb)

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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Jianyong Wu
    • 1
  • Yangcang Gong
    • 1
  • Minghui Cui
    • 1
  • Tingxiang Qi
    • 1
  • Liping Guo
    • 1
  • Jinfa Zhang
    • 2
  • Chaozhu Xing
    • 1
    Email author
  1. 1.Key Laboratory for Cotton Genetic Improvement, Ministry of Agriculture, Cotton Research InstituteChinese Academy of Agricultural SciencesAnyangChina
  2. 2.Department of Plant and Environmental SciencesNew Mexico State UniversityLas CrucesUSA

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