基于简化基因组开发青冈和滇青冈微卫星引物

欧阳泽怡; 李志辉; 牟虹霖; 姜小龙; 程勇; 吴际友

doi:10.12302/j.issn.1000-2006.202212002

欧阳泽怡, 李志辉, 牟虹霖, 姜小龙, 程勇, 吴际友

南京林业大学学报（自然科学版） ›› 2024, Vol. 48 ›› Issue (6) : 62-70.

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南京林业大学学报（自然科学版） ›› 2024, Vol. 48 ›› Issue (6) : 62-70. DOI: 10.12302/j.issn.1000-2006.202212002

研究论文

基于简化基因组开发青冈和滇青冈微卫星引物

欧阳泽怡 ¹^,² ,
李志辉 ¹ ,
牟虹霖 ¹ ,
姜小龙 ¹ ,
程勇 ³ ,
吴际友 ³^,^*

作者信息 +

Designing of microsatellite primers for Quercus glauca and Q. glaucoides (Fagaceae) based on RAD-seq data

OUYANG Zeyi ¹^,² ,
LI Zhihui ¹ ,
MOU Honglin ¹ ,
JIANG Xiaolong ¹ ,
CHENG Yong ³ ,
WU Jiyou ³^,^*

Author information +

文章历史 +

摘要

【目的】青冈(Quercus glauca)和滇青冈(Q. glaucoides)为珍贵树种,是东亚亚热带常绿阔叶林的重要建群种以及典型地理替代种,具有很高的生态和经济价值。开发其微卫星(simple sequence repeats,SSR)引物有助于分析其群体遗传格局及遗传多样性,为2种青冈林的管理和资源开发利用提供参考,也可为跨物种间的微卫星标记开发提供借鉴。【方法】分别基于3株青冈和3株滇青冈植株的简化基因组测序数据开发SSR引物。先后采用Stacks 2.0b软件process_radtags模型和SciRoKo 3.4软件对测序数据进行过滤和提取。利用Primer premier 6.0软件设计SSR引物。【结果】使用pyRAD 3.0.66软件对序列进行聚类,共鉴定出217个SSR位点,其中35%(76个)的SSR位点在青冈和滇青冈中均具有多态性。设计并筛选出28对SSR引物,分别在两个青冈群体和两个滇青冈群体(共48个个体)中进行巢式聚合酶链式反应(PCR)扩增。开发的28对SSR引物分布在青冈10条染色体上,SSR引物均在青冈和滇青冈个体中成功扩增,扩增率达到了90.7%。SSR分型分析结果表明,从遗传多样性方面看,所开发的28对引物中,共有176个等位基因;引物的等位基因数为3~13,平均为6.29个;期望杂合度为0.223~0.886,观察杂合度为 0.159~0.830。【结论】利用简化基因组数据可以快速、高效、经济地开发青冈和滇青冈通用微卫星引物,为后续2种青冈群体遗传学研究提供基础。也证明利用简化基因组测序数据,可开发近缘物种的通用微卫星引物。

Abstract

【Objective】Quercus glauca and Q. glaucoides are valuable and dominant species in the subtropical evergreen broadleaf forests of East Asia. They represent typical geographical vicarious species with significant ecological and economic importance. Therefore, the development of SSR primers for these two species can facilitate the analysis of genetic patterns and genetic diversity for the management and resource development of evergreen broadleaf forests as well as provide a reference for the development of microsatellite markers across species.【Method】This study developed SSR primers based on RADseq data from three Q. glauca and three Q. glaucoides individuals, respectively. The sequencing data were filtered and extracted using the process_radtags model in Stacks 2.0b software and SciRoKo 3.4 software sequentially. SSR primers were designed using Primer premier V6.0 software.【Result】The sequences were clustered using pyRAD 3.0.66, identifying a total of 217 SSR loci, 35% (76) of which were polymorphic in both species. Twenty-eight SSR primer pairs were designed and validated in two Q. glauca populations and two Q. glaucoides populations (48 individuals in total) through nested polymerase chain reaction (PCR) amplification. The 28 SSR primer pairs are distributed across ten chromosomes of the Q. glauca genome and successfully amplified in Q. glauca and Q. glaucoides individuals, with an amplification rate of 90.7%. The SSR genotyping analysis detected a total of 176 alleles, with the number of alleles per primer ranging from 3 to 13, and an average of 6.29. The expected and observed heterozygosity of the primers ranged from 0.223 to 0.886 and from 0.159 to 0.830, respectively.【Conclusion】The universal microsatellite primers developed in this study using RADseq data for Q. glauca and Q. glaucoides provide a basis for further population genetics studies of these species. In addition, this study demonstrates that RADseq data can be employed to rapidly, efficiently, and be used to cost-effectively develop universal microsatellite primers for closely related species.

导出引用

欧阳泽怡, 李志辉, 牟虹霖, 等. 基于简化基因组开发青冈和滇青冈微卫星引物[J]. 南京林业大学学报（自然科学版）. 2024, 48(6): 62-70 https://doi.org/10.12302/j.issn.1000-2006.202212002

OUYANG Zeyi, LI Zhihui, MOU Honglin, et al. Designing of microsatellite primers for Quercus glauca and Q. glaucoides (Fagaceae) based on RAD-seq data[J]. JOURNAL OF NANJING FORESTRY UNIVERSITY. 2024, 48(6): 62-70 https://doi.org/10.12302/j.issn.1000-2006.202212002

中图分类号： S795

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