基于简化基因组测序技术的甘薯HRM分子标记开发及其应用  

Development and Application of HRM Molecular Markers in Sweetpotato Based on Reduced-Representation Sequencing

作　　者：冯俊彦 郎涛[2] 张聪 李明[2] 青莉芳[2] 屈会娟 蒲志刚[2] 康乐[1] FENG Junyan;LANG Tao;ZHANG Cong;LI Ming;QING Lifang;QU Huijuan;PU Zhigang;KANG Le(College of Environmental Science and Engineering,China West Normal University,Nanchong,Sichuan 637002;Institute of Biotechnology and Nuclear Technology,Sichuan Academy of Agricultural Sciences,Chengdu,Sichuan 610061)

机构地区：[1]西华师范大学环境科学与工程学院,四川南充637002 [2]四川省农业科学院生物技术核技术研究所,四川成都610061

出　　处：《核农学报》2022年第7期1350-1361,共12页Journal of Nuclear Agricultural Sciences

基　　金：四川省科技计划项目(2019YFH0106);国家自然科学基金项目(31901588);四川省农业科学院学科建设推进工程项目计划(2021XKJS061);四川省科技支撑计划项目(2021YFY0027)。

摘　　要：目前甘薯中针对SNP位点的分子标记开发及应用的报道较少。为开发甘薯特异SNP分子标记,本研究利用简化基因组测序技术对甘薯种质材料进行测序,筛选稳定的SNP位点,将其开发为基于HRM技术的分子标记,并在甘薯种质材料中进行验证。通过对23个甘薯种质材料简化基因组测序数据的分析,共发现835 756个SNP多态性位点,筛选其中的3 650个含有SNP多态性位点的高质量测序片段,成功设计了134对引物;初步验证发现,22对(16.42%)引物没有扩增产物,15对(11.19%)引物扩增产物2个以上,36对(26.87%)引物的扩增产物没有差异。61对(45.52%)引物在8个样本中的扩增特异性好,而且样本之间有差异。最后筛选34对扩增多态性丰富的引物对52份甘薯种质材料进行扫描,发现材料间多态性介于27.27%~90.91%之间,平均多态性达到59.35%。聚类分析表明,参试52份甘薯种质材料之间的差异较小,多数材料与骨干亲本南瑞苕、徐薯18之间关系较近,国外引进甘薯材料和地方品种差异较大。建议在今后甘薯育种中尽量选用地方品种和国外甘薯品种,从而更好地拓宽甘薯遗传背景。本研究初步建立了基于简化基因组技术和HRM技术开发甘薯SNP分子标记的新思路,为今后甘薯SNP分子标记开发提供了参考。同时本研究开发的甘薯分子标记,丰富了甘薯分子标记类型,为甘薯研究者开展快速的分子标记研究提供了支持。There are few reports on the development and application of SNP markers in sweetpotato. In this study, we developed SNP molecular markers of HRM(high resolution melt) using sweetpotato reduced-representation sequencing data, and verified in sweetpotato germplasm. By analyzing sequence of 23 sweetpotato germplasm, a total of 835 756 SNP loci were found, 3 650 high-quality reads containing SNP polymorphic loci were selected, and 134 of primers were successfully developed. The results showed that 22 pairs of primers(16.42%) had no amplicons, and 15 primers(11.19%) had more than two amplified bands. The amplification products of 36 pairs of primers had no significant differences. 61 pairs of primers(45.52%) had specific amplification products, and differed among germplasm. Finally, 34 pairs of primers with rich amplification polymorphism were selected to amplify 52 sweetpotato germplasm. The results indicated that the polymorphism of molecular markers among germplasm ranged from 27.27% to 90.91%, and the average polymorphism was 59.35%. Cluster analysis showed that there were little differences among 52 sweetpotato accessions. Most bred varieties were close to the backbone parents Nancy Hall and Xushu 18. In contrast, there were great differences between the exotic sweetpotato germplasm and landraces. This study preliminarily established a new idea of developing sweetpotato SNP molecules markers based on reduced-representation sequencing technology and HRM technology, which provided a reference for the development of sweetpotato SNP molecular markers in the future. Furthermore, the molecular markers developed in this study enriched the types of molecular markers and provided support for sweetpotato rapid and efficient molecular marker study for sweet potato in the future.

关 键 词：甘薯 简化基因组测序 单核苷酸多态性 分子标记开发 

分 类 号：S531[农业科学—作物学]