Molecular Genetics and Genomics

, Volume 293, Issue 3, pp 623–633 | Cite as

Gene-based SNP identification and validation in soybean using next-generation transcriptome sequencing

  • Yong Guo
  • Bohong Su
  • Junyong Tang
  • Fulai Zhou
  • Li-Juan Qiu
Original Article


Gene-based molecular markers are increasingly used in crop breeding programs for marker-assisted selection. However, identification of genetic variants associated with important agronomic traits has remained a difficult task in soybean. RNA-Seq provides an efficient way, other than assessing global expression variations of coding genes, to discover gene-based SNPs at the whole genome level. In this study, RNA isolated from four soybean accessions each with three replications was subjected to high-throughput sequencing and a range of 44.2–65.9 million paired-end reads were generated for each library. A total of 75,209 SNPs were identified among different genotypes after combination of replications, 89.1% of which were located in expressed regions and 27.0% resulted in amino acid changes. GO enrichment analysis revealed that most significant enriched genes with nonsynonymous SNPs were involved in ribonucleotide binding or catalytic activity. Of 22 SNPs subjected to PCR amplification and Sanger sequencing, all of them were validated. To test the utility of identified SNPs, these validated SNPs were also assessed by genotyping a relative large population with 393 wild and cultivated soybean accessions. These SNPs identified by RNA-Seq provide a useful resource for genetic and genomic studies of soybean. Moreover, the collection of nonsynonymous SNPs annotated with their predicted functional effects also provides a valuable asset for further discovery of genes, identification of gene variants, and development of functional markers.


RNA-Seq Single-nucleotide polymorphism Soybean Nonsynonymous SNPs Next-generation sequencing 



This work was supported by the National Natural Science Foundation of China (31471520), 13th Five-Year Plan for Precise Identification and Germplasm Enhancement of Economic Crops, and the Agricultural Science and Technology Innovation Program (ASTIP) of Chinese Academy of Agricultural Sciences.

Author contributions

YG and LJQ conceived and designed the experiments. YG, BS, JT and FZ performed the experiments. YG and LJQ analyzed data and wrote the manuscript. All authors read and approved the manuscript.

Compliance with ethical standards

Conflict of interest

All authors declare that they have no conflict of interest.

Research involving human and animal participants

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

Supplementary material

438_2017_1410_MOESM1_ESM.xlsx (9 kb)
Online Resource 1. The morphology of soybean accessions used for RNA-seq (XLSX 9 KB)
438_2017_1410_MOESM2_ESM.xlsx (9 kb)
Online Resource 2. Primers list for SNP validation (XLSX 9 KB)
438_2017_1410_MOESM3_ESM.xlsx (29 kb)
Online Resource 3. The information of soybean accessions used for genotyping (XLSX 28 KB)
438_2017_1410_MOESM4_ESM.xlsx (3.1 mb)
Online Resource 4. All SNPs identified in the soybean transcriptome sequencing (XLSX 3161 KB)


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2017

Authors and Affiliations

  • Yong Guo
    • 1
  • Bohong Su
    • 1
    • 2
  • Junyong Tang
    • 1
  • Fulai Zhou
    • 1
  • Li-Juan Qiu
    • 1
  1. 1.The National Key Facility for Crop Gene Resources and Genetic Improvement (NFCRI) and MOA Key Labs of Crop Germplasm and Soybean Biology (Beijing), Institute of Crop ScienceChinese Academy of Agricultural SciencesBeijingPeople’s Republic of China
  2. 2.College of AgricultureNortheast Agricultural UniversityHarbinPeople’s Republic of China

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