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

Abstract

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.

Population transcriptomic sequencing reveals allopatric divergence and local adaptation in Pseudotaxus chienii (Taxaceae)

“…We used RNA-Seq, a commonly used NGS approach, to quantify the expression level of each unigene in each individual by mapping clean reads to reference sequences. SNP markers, third-generation molecular markers, have wide applicability, particularly SNP markers from transcriptome sequences, which can effectively reveal functional SNPs at the whole-genome level [28]. In this study, 13,545 high-quality SNPs were identified in 108 individuals of P. chienii based on transcriptome data to explore the driving mechanism of this species' adaptations to its natural habitat.…”

Advances in Transcriptome Analyses Using RNA Sequencing Technology in Soybean Plants [Glycine max]

“…This study represents the first report of a comprehensive transcriptomic and proteomic analysis of soybean sprout in response to supplemental Ca 2+ . Guo et al (2018) reported that RNA-seq data generated from four soybean accessions. A total of 75 209 SNPs were identified, 89.1% of which were located in expressed regions and 27.0% resulted in amino acid changes.…”

SNP Detection in Pinus pinaster Transcriptome and Association with Resistance to Pinewood Nematode

“…With the rise of next generation sequencing, the developing of molecular markers has become easier and more affordable, even for non-model species [12]. RNA-seq is one of these technologies that has been frequently used for the discovery of molecular markers, such as single nucleotide polymorphisms (SNPs) and simple-sequence repeats (SSRs) [15][16][17]. As RNA-seq produces information mainly on protein coding regions, polymorphisms associated with phenotype are more easily linked to a functional effect.…”