Genes & Genomics

, Volume 39, Issue 10, pp 1117–1127 | Cite as

Transcriptome comparative analysis between the cytoplasmic male sterile line and fertile line in soybean (Glycine max (L.) Merr.)

  • Jiajia Li
  • Shouping Yang
  • Junyi Gai
Research Article


To further elucidate the molecular mechanism and fertility restoration of cytoplasmic male sterility (CMS) in soybean, a comparative transcriptome analysis was conducted between the CMS line NJCMS1A, restorer line NJCMS1C and their hybrid F1 progeny (NJCMS1A × NJCMS1C) using RNA-Seq strategy. After pairwise comparative analysis of these soybean lines, 294, 222, and 288 differentially expressed genes (DEGs) were identified, respectively. Further bioinformatic analysis indicated that these DEGs were involved in diverse molecular functions and metabolic pathways. qRT-PCR analysis validated that the gene expression pattern in RNA-Seq was reliable. These results significantly showed that the male sterility and fertility restoration in NJCMS1A might be related to a series of the abnormal of growth development and metabolic processes, such as pollen development, DNA methylation process, pollen viability, cell wall development, programmed cell death, as well as carbohydrate and energy metabolism. This study could facilitate our understanding of the molecular mechanisms and fertility restoration behind CMS in soybean.


Soybean (Glycine max (L.) Merr.) Cytoplasmic male sterility (CMS) Fertility restoration Transcriptomics Differentially expressed genes (DEGs) 



This work was supported by the National Key Research and Development Program of China (2016YFD0101500, 2016YFD0101504), the National Hightech R & D Program of China (2011AA10A105), and the Program for Changjiang Scholars and Innovative Research Team in University (PCSIRT13073).

Author contributions

SPY and JYG conceived this study. JJL and SPY designed the experimental plan and drafted and revised the manuscript. JJL analyzed and interpreted the sequence data. All authors read and approved the final manuscript.

Compliance with ethical standards

Conflict of interest

All the authors declare that they have no conflict of interest.

Ethical approval

The article does not contain any studies with human subjects or animals performed by any of the authors.

Supplementary material

13258_2017_578_MOESM1_ESM.xls (234 kb)
Table S1: Number of differentially expressed genes (DEGs) between different comparison groups. Table S1-1: Number of DEGs between NJCMS1A and NJCMS1C. Table S1-2: Number of DEGs between NJCMS1A and F1. Table S1-3: Number of DEGs between NJCMS1C and F1. (XLS 233 KB)
13258_2017_578_MOESM2_ESM.xls (36 kb)
Table S2: KEGG pathways enriched of differentially expressed gene (DEGs) between different compared groups. Table S2-1: KEGG pathways enriched of DEGs between NJCMS1A and NJCMS1C. Table S2-2: KEGG pathways enriched of DEGs between NJCMS1A and F1. Table S2-3: KEGG pathways enriched of DEGs between NJCMS1C and F1. (XLS 36 KB)
13258_2017_578_MOESM3_ESM.xls (32 kb)
Table S3: Comparison of expression patterns between RNA-Seq and qRT-PCR. Table S3-1: DEGs confirmed by qRT-PCR using the same sample as that in RNA-Seq. Table S3-2: DEGs confirmed by qRT-PCR using different sample from that in RNA-Seq. Table S3-3: The compare results between qRT-PCR and RNA-Seq of DEG (Glyma.16G045100) shared in three different compared groups. (XLS 32 KB)


  1. An H, Yang ZH, Yi B, Wen J, Shen JX, Tu JX, Ma CZ, Fu TD (2014) Comparative transcript profiling of the fertile and sterile flower buds of pol CMS in B. napus. BMC Genomics 15:258CrossRefPubMedPubMedCentralGoogle Scholar
  2. Ansorge WJ (2009) Next-generation DNA sequencing techniques. New Biotechnol 25:195–203CrossRefGoogle Scholar
  3. Benjamini BY, Hochberg Y (1995) Controlling the false discovery rate: a practical and powerful approach to multiple testing. J R Stat Soc 57:289–300Google Scholar
  4. Benjamini BY, Yekutieli D (2001) The control of the false discovery rate in multiple testing under dependency. Ann Stat 29:1165–1188CrossRefGoogle Scholar
  5. Conesa A, Götz S, García-Gómez JM, Terol J, Talón M, Robles M (2005) Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics 21:3674–3676CrossRefPubMedGoogle Scholar
  6. Ding DR, Gai JY, Cui ZL, Yang SP, Qiu JX (1999) Development and verification of the cytoplasmic-nuclear male sterile soybean line NJCMS1A and its maintainer NJCMS1B. Chin Sci Bull 44:191–192CrossRefGoogle Scholar
  7. Ding DR, Gai JY, Cui ZL, Qiu JX (2002) Development of a cytoplasmic-nuclear male-sterile line of soybean. Euphytica 124:85–91CrossRefGoogle Scholar
  8. Fan JM (2003) Studies on cyto-morphological and cyto-chemical features of cytoplasmic-nuclear male-sterile lines of soybeans (Glycine max (L.) Merr.). M. Sc. Thesis, Nanjing Agricultural University, Supervisor: S.P. YangGoogle Scholar
  9. Feldmann KA (2001) Cytochrome P450s as genes for crop improvement. Curr Opin Plant Biol 4:162–167CrossRefPubMedGoogle Scholar
  10. Fry SC, Smith RC, Renwick KF, Martin DJ, Hodge S, Matthews KJ (1992) Xyloglucan endotransglycosylase, a new wall-loosening enzyme activity from plants. Biochem J 282:821–828CrossRefPubMedPubMedCentralGoogle Scholar
  11. Gai JY, Cui ZL, Ji DF, Ren ZJ, Ding DR (1995) A report on the nuclear cytoplasmic male sterility from a cross between two soybean cultivars. Soy Genet Newsl 22:55–58Google Scholar
  12. Ge X, Dietrich C, Matsuno M, Li G, Berg H, Xia Y (2005) An Arabidopsis aspartic protease functions as an anti-cell-death component in reproduction and embryogenesis. EMBO Rep 6:282–288CrossRefPubMedPubMedCentralGoogle Scholar
  13. Havey MJ (2004) The use of cytoplasmic male sterility for hybrid seed production. In: Daniell H, Chase CD (eds) Molecular biology and biotechnology of plant organelles. Springer, Dordrecht, pp 623–634CrossRefGoogle Scholar
  14. Huang F, Xu GL, Chi YJ, Liu HC, Xue Q, Zhao TJ, Gai JY, Yu DY (2014) A soybean MADS-box protein modulates floral organ numbers, petal identity and sterility. BMC Plant Biol 14:89CrossRefPubMedPubMedCentralGoogle Scholar
  15. Kurasawa K, Matsui A, Yokoyama R, Kuriyama T, Yoshizumi T, Matsui M, Suwabe K, Watanabe M, Nishitani K (2009) The AtXTH28 gene, a xyloglucan endotransglucosylase/hydrolase, is involved in automatic self-pollination in Arabidopsis thaliana. Plant Cell Physiol 50:413–422CrossRefPubMedPubMedCentralGoogle Scholar
  16. Li JJ, Han SH, Ding XL, He TT, Dai JY, Yang SP, Gai JY (2015) Comparative transcriptome analysis between the cytoplasmic male sterile line NJCMS1A and its maintainer NJCMS1B in soybean (Glycine max (L.) Merr.). PLoS ONE 10:e0126771CrossRefPubMedPubMedCentralGoogle Scholar
  17. Liu C, Ma N, Wang PY, Fu N, Shen HL (2013) Transcriptome sequencing and de novo analysis of a cytoplasmic male sterile line and its near-isogenic restorer line in chili pepper (Capsicum annuum L.). PLoS ONE 8:e65209CrossRefPubMedPubMedCentralGoogle Scholar
  18. Mamun EA, Alfred S, Cantrill LC, Overall RL, Sutton BG (2006) Effects of chilling on male gametophyte development in rice. Cell Biol Int 30:583–591CrossRefPubMedGoogle Scholar
  19. Ng HH, Adrian B (1999) DNA methylation and chromatin modification. Curr Opin Genet Dev 9:158–163CrossRefPubMedGoogle Scholar
  20. Nishitani K, Tominaga R (1992) Endo-xyloglucan transferase, a novel class of glycosyltransferase that catalyzes transfer of a segment of xyloglucan molecule to another xyloglucan molecule. J Biol Chem 267:21058–21064PubMedGoogle Scholar
  21. O’Keefe DP, Tepperman JM, Dean C, Leto KJ, Erbes DL, Odell JT (1994) Plant expression of a bacterial cytochrome P450 that catalyses activation of a sulfonylurea pro-herbicide. Plant Physiol 105:473–482CrossRefPubMedPubMedCentralGoogle Scholar
  22. Rose JKC, Braam J, Fry SC, Nishitani K (2002) The XTH family of enzymes involved in xyloglucan endotransglucosylation and endohydrolysis: current perspectives and a new unifying nomenclature. Plant Cell Physiol 43:1421–1435CrossRefPubMedGoogle Scholar
  23. Schuster SC (2008) Next-generation sequencing transforms today’s biology. Nat Methods 5:16–18CrossRefPubMedGoogle Scholar
  24. Sheoran IS, Sawhney VK (2010) Proteome analysis of the normal and Ogura (ogu) CMS anthers of Brassica napus to identify proteins associated with male sterility. Botany 88:217–230CrossRefGoogle Scholar
  25. Stefan M, Anja P, Ulrich M (2010) Multifunctional flavonoid dioxygenases: flavonol and anthocyanin biosynthesis in Arabidopsis thaliana L. Phytochemistry 71:1040–1049CrossRefGoogle Scholar
  26. Tang HB, Wang XY, Bowers JE, Ming R, Alam M, Paterson AH (2008) Unraveling ancient hexaploidy through multiply-aligned angiosperm gene maps. Genome Res 18:1944–1954CrossRefPubMedPubMedCentralGoogle Scholar
  27. Trapnell C, Pachter L, Salzberg SL (2009) TopHat: discovering splice junctions with RNA-SEq. Bioinformatics 25:1105–1111CrossRefPubMedPubMedCentralGoogle Scholar
  28. Trapnell C, Williams BA, Pertea G, Mortazavi A, Kwan G, van Baren MJ, Salzberg SL, Wold BJ, Pachter L (2010) Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat Biotechnol 28:511–515CrossRefPubMedPubMedCentralGoogle Scholar
  29. Trapnell C, Roberts A, Goff L, Pertea G, Kim D, Kelley DR, Pimentel H, Salzberg SL, Rinn JL, Pachter L (2012) Differential gene and transcript expression analysis of RNA-Seq experiments with TopHat and Cufflinks. Nat Protoc 7:562–578CrossRefPubMedPubMedCentralGoogle Scholar
  30. Trapnell C, Hendrickson DG, Sauvageau M, Goff L, Rinn JL, Pachter L (2013) Differential analysis of gene regulation at transcript resolution with RNA-SEq. Nat Biotechnol 31:46–53CrossRefPubMedGoogle Scholar
  31. Van der Meer IM, Stam ME, Tunen AJ, Mol JN, Suitje AR (1992) Antisense inhibition of flavonoid biosynthesis in petunia anthers results in male sterility. Plant Cell 4:253–262CrossRefPubMedPubMedCentralGoogle Scholar
  32. Wang LG, Wang SQ, Li W (2012) RSeQC: quality control of RNA-Seq experiments. Bioinformatics 28:2184–2185CrossRefPubMedGoogle Scholar
  33. Wei MM, Song MZ, Fan SL, Yu SX (2013) Transcriptomic analysis of differentially expressed genes during anther development in genetic male sterile and wild type cotton by digital gene-expression profiling. BMC Genomics 14:97CrossRefPubMedPubMedCentralGoogle Scholar
  34. Werck-Reichhart D, Hehn A, Didierjean L (2000) Cytochromes P450 for engineering herbicide tolerance. Trends Plant Sci 5:116–123CrossRefPubMedGoogle Scholar
  35. Wu ZM, Cheng JW, Qin C, Hu ZQ, Yin CX, Hu KL (2013) Differential proteomic analysis of anthers between cytoplasmic male sterile and maintainer lines in Capsicum annuum L. Int J Mol Sci 14:22982–22996CrossRefPubMedPubMedCentralGoogle Scholar
  36. Xie C, Mao XZ, Huang JJ, Ding Y, Wu JM, Dong S, Kong L, Gao G, Li CY, Wei L (2011) KOBAS 2.0: a web server for annotation and identification of enriched pathways and diseases. Nucleic Acids Res 39:W316–W322CrossRefPubMedPubMedCentralGoogle Scholar
  37. Xu PZ, Yan WG, He J, Li Y, Zhang HY, Peng H, Wu XJ (2013) DNA methylation affected by male sterile cytoplasm in rice (Oryza sativa L.). Mol Breed 31:719–727CrossRefGoogle Scholar
  38. Yan XH, Dong CH, Yu JY, Liu WH, Jiang CH, Liu J, Hu Q, Fang XP, Wei WH (2013) Transcriptome profile analysis of young floral buds of fertile and sterile plants from the self-pollinated offspring of the hybrid between novel restorer line NR1 and Nsa CMS line in Brassica napus. BMC Genomics 14:26CrossRefPubMedPubMedCentralGoogle Scholar
  39. Yang SP, Duan MP, Meng QC, Qiu JX, Fan JM, Zhao TJ, Yu DY, Gai JY (2007) Inheritance and gene tagging of male fertility restoration of cytoplasmic-nuclear male-sterile line NJCMS1A in soybean. Plant Breed 126:302–305CrossRefGoogle Scholar
  40. Zenoni S, Ferrarini A, Giacomelli E, Xumerle L, Fasoli M, Malerba G, Bellin D, Pezzotti M, Delledonne M (2010) Characterization of transcriptional complexity during berry development in Vitis vinifera using RNA-SEq. Plant Physiol 152:1787–1795CrossRefPubMedPubMedCentralGoogle Scholar
  41. Zheng R, Yue SY, Xu XY, Liu JY, Xu Q, Wang XL, Han L, Yu DY (2012) Proteome analysis of the wild and YX-1 male sterile mutant anthers of wolfberry (Lycium barbarum L.). PLoS ONE 7:e41861CrossRefPubMedPubMedCentralGoogle Scholar
  42. Zhu J, Yang ZN (2013) The research progress of pollen wall development. Chin J Nat 35:112–117 (in Chinese) Google Scholar

Copyright information

© The Genetics Society of Korea and Springer Science+Business Media B.V. 2017

Authors and Affiliations

  1. 1.Soybean Research Institute, National Center for Soybean Improvement, Key Laboratory of Biology and Genetic Improvement of Soybean (General, Ministry of Agriculture), State Key Laboratory of Crop Genetics and Germplasm Enhancement, Jiangsu Collaborative Innovation Center for Modern Crop ProductionNanjing Agricultural UniversityNanjingChina
  2. 2.College of AgronomyAnhui Agricultural UniversityHefeiChina

Personalised recommendations