基于SLAF-seq技术的舒玛栎群体遗传多样性与遗传结构分析

【目的】对美国引进的不同种源舒玛栎群体进行遗传多样性与遗传结构分析,揭示其遗传分化特点及单株间遗传关系,为舒玛栎种质资源的保护与品种选育提供理论依据。【方法】以舒玛栎6个种源30个单株以及外类群纳塔栎5个单株为材料,基于SLAF-seq技术进行简化基因组测序,开发一批SNP标记并选择其中多态性的SNP标记进行基因分型。利用GenAlex、Arlequin、MEGA、Admixture和Cluster等软件进行遗传多样性参数估算、F统计量及分子分差分析、进化树构建、遗传结构与PCA主成分分析。【结果】35个栎树个体SLAF测序平均深度11×,碱基质量(Q₃₀)平均为93%,GC含量平均为38.9%。共获得4 256 436个SLAF标签,开发多态性SNP标记8 459 025个,SNP完整度平均为79.29%,杂合率平均为14.15%。舒玛栎6个种源平均有效等位基因数为1.31个,多态性位点比例平均为49.21%;观测杂合度(H_o)与期望杂合度(H_e)变化范围分别为0.13~0.16和0.17~0.21;多态信息含量(PIC)、香农指数(I)、Nei’s基因多样性指数(H)和群体内的近交系数(F_IS)平均值分别为0.15、0.34、0.09和0.19。不同种源间Nei’s遗传距离和遗传分化系数(F_ST)变化范围为0.08~0.18和0.15~0.39。分子方差分析表明舒玛栎遗传变异主要来自个体间。遗传结构分析显示30个舒玛栎个体来源于3个原始的祖先。【结论】舒玛栎群体遗传多样性水平较高,群体间遗传分化程度较大,在品种选育中应注重群体内个体优树的选择。

Genetic diversity and structure analyses of Quercus shumardii populations based on SLAF-seq technology

【Objective】Oaks are the most important productive and dominant species in broadleaf forests in the subtropics and temperate zones, and they play irreplaceable roles in forest ecology. As a precious and colorful broadleaf tree species, Quercus shumardii has been extensively utilized for commercial timber production and landscape planting. To provide a theoretical basis for the germplasm protection and variety selection, the genetic diversity and genetic structure of different provenances of Q. shumardii introduced from the USA were evaluated to reveal the characteristics of genetic differentiation and genetic relationships among individuals. 【Method】Thirty-five Quercus individuals were sampled, including 30 individuals from six provenances of Q. shumardii and five individuals of Q. nuttallii as an outgroup. A set of SNP markers was developed based on SLAF sequencing according to a non-reference genome protocol, of which the polymorphic SNP markers with a minor allele frequency (MAF) > 0.05 and miss rate < 0.2 were screened and used for a genotyping. Several software programs such as GenAlex, Arlequin, MEGA, Admixture and Cluster were used for genetic diversity parameter estimation, F statistics and molecular variance analysis, phylogenetic tree construction, genetic structure analysis, and principal component analysis, respectively. 【Result】Average sequencing depth of the 35 Quercus individuals was 11-fold. The values of Q₃₀ and GC content of the seven provenances ranged from 90.24% to 95.12% and from 38.49% to 39.35%, with average values of 93% and 38.9%, respectively. After assembly, 4 256 436 SLAF tags were obtained, and 8 459 025 polymorphic SNP markers were identified, of which the average integrity and the heterozygosity ratio of SNPs was 79.29% and 14.15%, respectively. Among the six different provenances of Q. shumardii, the average effective number of alleles was 1.31, and the percentage of polymorphic loci was 49.21%. The observed and expected heterozygosity (H_o and H_e) ranged from 0.13 to 0.16 (mean 0.15) and 0.17 to 0.21 (mean 0.19), respectively. Average values of polymorphism information content (PIC), Shannon’s information index (I), Nei’s gene diversity (H), and inbreeding coefficient (${F_I}_S$) were 0.15, 0.34, 0.09 and 0.19, respectively. The pairwise genetic differentiation index (F_ST) and genetic distance among provenances of Q. shumardii ranged from 0.08 to 0.18 and from 0.15 to 0.39, respectively. The molecular variance analysis demonstrated that the differentiation among individuals contributed 84.88% of the total variation, and the remaining 15.12% was among populations. Furthermore, the genetic structure results showed that 30 individuals of Q. shumardii originated from three primitive ancestors, of which two groups contained TX and MS provenances, respectively, and the other group contained LA, MO, OH and PA provenances. The cluster analysis demonstrated that all 35 individuals were separated into two clades, i.e., the species Q. shumardii and Q. nuttallii, which belonged to the traditional taxonomy of Quercus. 【Conclusion】A relatively high level of genetic diversity and genetic differentiation was detected in the Q. shumardii population, suggesting that the superior individual tree among populations should be focused on for the variety selection.