Gene-based SNP identification and validation in soybean using next-generation transcriptome sequencing
Gene-based SNP identification and validation in soybean using next-generation transcriptome sequencing
复制标题
使用下一代转录组测序对大豆进行基于基因的 SNP 鉴定和验证
DOI：
10.1007/s00438-017-1410-5
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发表时间：
2018-06-01
影响因子：
3.1
通讯作者：
Qiu, Li-Juan
中科院分区：
文献类型：
--
作者：
Guo, Yong;Su, Bohong;Qiu, Li-Juan
基于基因的分子标记越来越多地用于作物育种计划中的标记辅助选择。然而，鉴定与重要农艺性状相关的遗传变异在大豆中仍然是一项困难的任务。RNA-Seq提供了一种有效的方法，而不是评估编码基因的全局表达变异，以发现基于基因的SNP在整个基因组水平。在这项研究中，从四个大豆种质中分离的RNA进行高通量测序，每个文库产生4420万至6590万个配对末端读段。在不同基因型间共检测到75，209个SNPs，其中89.1%位于表达区，27.0%导致氨基酸改变。GO富集分析显示，最显着的富集基因与非同义SNP参与核糖核苷酸结合或催化活性。对22个SNPs进行PCR扩增和桑格测序，均得到验证。为了测试所鉴定的SNP的效用，还通过对具有393个野生和栽培大豆种质的相对大的群体进行基因分型来评估这些经验证的SNP。通过RNA-Seq技术鉴定的这些SNPs为大豆的遗传和基因组研究提供了有用的资源。此外，收集的非同义SNP注释其预测的功能效果也提供了一个有价值的资产，进一步发现的基因，基因变异的鉴定，和功能标记的发展。
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.