Abstract
Key message
Based on Ginkgo biloba transcriptome data, 22 SNP loci using high-resolution melting curve technology were validated and genetic diversity was analyzed in three populations.
Abstract
Ginkgo biloba (Ginkgo) is a long-lived dioecious gymnosperm with unique morphological characteristics and play a significant role in evolutionary relationship research. In this study, we used Illumina paired-end RNA sequencing technology for facilitating the gene discovery and single nucleotide polymorphism (SNP) development in Ginkgo. We collected 44.08 G clean bases from six transcriptome datasets. The transcriptome data generated 98,919 unigenes among which 42,667 (43.13%) were successfully annotated. A total of 139,854 putative SNPs were identified; most of the SNPs were transition-type with the nucleotide transitions C–T or A–G. Further, 54532 (38.99%) SNPs were found in protein-coding regions: 23483 (43.06%) were synonymous and 31049 (56.94%) were nonsynonymous. 22 SNPs were subjected to PCR amplification and Sanger sequencing, and all of them were validated. To test the practicability of identified SNPs, these validated SNPs were also assessed by genotyping three natural populations with 84 individuals by high-resolution melting curve (HRM) analysis. Observed and expected heterozygosity varied from 0.0119 to 0.9643 and from 0.0581 to 0.5024, respectively. HRM technology was first applied for the SNP genotyping in Ginkgo. SNPs identified by RNA-Seq provided a useful resource for genetic and genomic studies in Ginkgo. Moreover, the collection of nonsynonymous SNPs annotated with their predicted functional effects also provided a valuable asset for further discovery of genes, identification of gene variants, and development of functional markers.
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Acknowledgements
This study was supported by the Special Fund for Forest Scientific Research in the Public Welfare (201504105), the Agricultural Science and Technology Independent Innovation Funds of Jiangsu Province (CX(16)1005), the National Key Research and Development Program of China (2017YFD0600700), Postgraduate Research & Practice Innovation Program of Jiangsu Province (KYCX18_0954), and the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).
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Supplementary Table S1
Unigenes were annotated to the seven databases (DOCX 14 KB)
Supplementary Figure S1
Gene ontology (GO) classification of unigenes. All the annotated unigenes were divided into three functional GO categories: biological process (BP), cellular component (CC) and molecular function (MF) (PDF 6 KB)
Supplementary Figure S2
EuKaryotic Orthologous Groups (KOG) annotation of putative proteins. The x-axis indicates the names of the 26 groups of KOG. The y-axis indicates the number of genes annotated to the group out of the total number of genes annotated (PDF 5 KB)
Supplementary Figure S3
The several typical SNP genotypes by Sanger sequencing. From A to H, the loci with the presence of a bimodal peak was identified to be heterozygous (TIF 17698 KB)
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Wu, Y., Zhou, Q., Huang, S. et al. SNP development and diversity analysis for Ginkgo biloba based on transcriptome sequencing. Trees 33, 587–597 (2019). https://doi.org/10.1007/s00468-018-1803-z
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DOI: https://doi.org/10.1007/s00468-018-1803-z