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

OUYANG Zeyi, LI Zhihui, MOU Honglin, JIANG Xiaolong, CHENG Yong, WU Jiyou

JOURNAL OF NANJING FORESTRY UNIVERSITY ›› 2024, Vol. 48 ›› Issue (6) : 62-70.

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

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

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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.

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Quercus glauca / Quercus glaucoides / microsatellite / RAD-seq

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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

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