3 Biotech

, 9:22 | Cite as

Exploration of miRNA-mediated fertility regulation network of cytoplasmic male sterility during flower bud development in soybean

  • Xianlong Ding
  • Hao Zhang
  • Hui Ruan
  • Yanwei Li
  • Linfeng Chen
  • Tanliu Wang
  • Ling Jin
  • Xiaoqiang Li
  • Shouping YangEmail author
  • Junyi GaiEmail author
Original Article


Cytoplasmic male sterility (CMS) plays an important role in the production of soybean hybrid seeds. MicroRNAs (miRNAs) are a class of non-coding endogenous ~ 21 nt small RNAs that play crucial roles in flower and pollen development by targeting genes in plants. To dissect the function of miRNAs in soybean CMS, a total of 558 known miRNAs, 10 novel miRNAs, and 466 target genes were identified in flower buds of the soybean CMS line NJCMS1A and its restorer line NJCMS1C through small RNA sequencing and degradome analysis. In addition, miRNA-mediated editing events were also observed, and the two most frequently observed editing types (A → G and C → U) were validated by cloning and sequencing. And as the base editing occurred, some targets were filtered, such as gma-miR2118b-P6GT with Glyma.08G122000.2. Further integrated analysis of transcriptome and small RNA found some miRNAs and their targets’ expression patterns showing a negative correlation, such as gma-miR156b/GmSPL9a and gma-miR4413b/GmPPR. Furthermore, opposite expression pattern was observed between gma-miR156b and GmSPL9 during early stage of flower bud development. Taken together, the regulatory network of gma-miR156b/GmSPL9 and gma-miR4413b/GmPPR with flower bud development in soybean CMS was developed. Most importantly, previous reports showed that these targets might be related to pollen development and male sterility, suggesting that both conserved and species-specific miRNAs might act as regulators of flower bud development in soybean CMS. These findings may provide a better understanding of the miRNA-mediated regulatory networks of CMS mechanisms in soybean.


Soybean (Glycine max (L.) Merr.) Cytoplasmic male sterility Fertility regulation network miRNA Target gene 



Cytoplasmic male sterility


Minimum free energies


Minimal folding energy indices




Pentatricopeptide repeat-containing


Quantitative real time PCR


Restorer of fertility


Squamosa promoter-binding protein-like



This work was supported by grants from the National Key R&D 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 (PCSIRT_17R55, PCSIRT13073).

Author Contributions

SPY, JYG and XLD conceived and designed the experiments. XLD, HZ, HR, YWL, LFC, TLW, LJ and XQL performed the experiments. XLD analyzed the data. XLD, HZ and HR contributed reagents/materials/analysis tools. XLD, YWL, LFC, TLW, LJ and XQL conceived the qRT-PCR experiments and analyzed the data. XLD, SPY conceived the experiments and wrote the manuscript. All authors read and approved the final manuscript.

Compliance with ethical standards

Conflict of interest

All the authors declare that they do not have conflict of interest.

Ethical approval

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

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Copyright information

© King Abdulaziz City for Science and Technology 2019

Authors and Affiliations

  • Xianlong Ding
    • 1
    • 2
  • Hao Zhang
    • 1
  • Hui Ruan
    • 1
  • Yanwei Li
    • 1
  • Linfeng Chen
    • 1
  • Tanliu Wang
    • 1
  • Ling Jin
    • 1
  • Xiaoqiang Li
    • 1
  • Shouping Yang
    • 1
    Email author
  • Junyi Gai
    • 1
    Email author
  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 Production, College of AgricultureNanjing Agricultural UniversityNanjingChina
  2. 2.College of Life SciencesNanjing Agricultural UniversityNanjingChina

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