基于SSR分子标记的地被菊遗传多样性分析和指纹图谱构建

Genetic diversity analysis and fingerprints of Chrysanthemum×morifolium based on SSR molecular markers

摘要:

为了从分子水平上研究地被菊（Chrysanthemum×morifolium）种质资源的遗传多样性并建立地被菊品种的指纹图谱库，筛选出多态性高的引物用于地被菊品种间鉴定、亲缘关系分析和分子标记辅助选种体系的建立，本研究利用多态性好、条带清晰、重复性好的12对引物对91份地被菊材料和14份菊属近缘种材料进行简单重复序列（simple sequence repeat，SSR）分子标记和遗传多样性分析，从12对引物中筛选出9对核心引物对受试材料进行指纹图谱构建。结果显示，在105个样品中，12对引物检测出104个等位位点，范围为2—26，平均每个点位检测出9.25个等位基因数，平均每个点位检测得到的有效等位基因数（number of effective alleles，N_e）为2.745 6，范围为1.276 0—4.742 5；Shannon信息指数（Shannon genetic diversity index，I）变化范围是0.513 3—2.239 9，均值是1.209 0；Nei’s基因多样性指数（Nei’s gene diversity index，H）范围是0.216 3—0.789 1，均值是0.578 0；观测杂合度（observed heterozygosity，H_o）的范围是0.223 3—0.895 2，均值是0.557 5；期望杂合度（expected heterozygosity，H_e）的范围是0.217 4—0.793 3，均值是0.580 8；多态信息含量（polymorphism information content，PIC）值变化范围是0.211 5—0.774 0，均值是0.532 9；遗传相似性系数（genetic similarity，GS）范围为0.228 5—1.000 0，均值是0.608 3。聚类分析表明，在遗传距离（genetic distance，GD）=0.30时，受试材料可以分为两个类群。Structure群体结构分析将受试材料分为3个种群和1个混合种群。从12对引物中筛选出可完全区分105份受试材料的9对核心引物，构建了91份地被菊材料和14份菊属近缘种材料的指纹图谱。地被菊材料之间具有显著的遗传差异和丰富的遗传多样性，对于地被菊的园林应用和品种选育具有重要意义。地被菊品种和菊属近缘种的指纹图谱库的构建，一定程度上揭示了105份实验材料的亲缘关系，为今后地被菊品种鉴定与筛选体系的研究提供了技术支撑。

Abstract:

The present study aims to explore the genetic diversity of germplasm resources of Chrysanthemum×morifolium (hereinafter, C.×morifolium) at the molecular level and to establish a fingerprint database of C.×morifolium varieties. We employed 12 pairs of primers with high levels of polymorphism, clear bands, and high degrees of reproducibility to analyze the SSR molecular markers and genetic diversity of 91 C.×morifolium materials and 14 chrysanthemum- related materials. With regard to constructing the fingerprints of the tested materials, we chose 9 pairs of core primers. The findings revealed that 12 primer pairs detected 104 alleles in 105 samples, ranging from 2 to 26. The average number of observed alleles (N_a) per site was 9.25. The average number of effective alleles (N_e) per site was 2.745 6, with its range being 1.276 0 to 4.742 5. Shannon genetic diversity index (I) values ranged between 0.513 3 and 2.239 9 (M=1.209 0). Nei's gene diversity index (H) ranged between 0.216 3 and 0.789 1 (M=0.578 0). The observed heterozygosity (H_o) ranged between 0.223 3 and 0.895 2 (M=0.557 5). The expected heterozygosity (H_e) ranged between 0.217 4 and 0.793 3 (M=0.580 8). The polymorphism information content (PIC) ranged between 0.211 5 and 0.774 0 (M=0.532 9). The genetic similarity (GS) ranged between 0.228 5 and 1.000 0 (M=0.608 3). Cluster analysis revealed that when the genetic distance (GD) equals to 0.30, the tested materials can be classified into 2 groups. When the GD equals to 0.27, the first group can be divided into 6 subgroups; accordingly, 105 tested materials can be divided into 7 subgroups. The cophenetic correlation test was carried out based on the cluster analysis, and the corresponding results showed that the cluster map correlated with the genetic similarity coefficient (r=0.952 73). According to the results of Structure population analysis, we obtained the optimal population number, with the true number of populations (K) being 3 and the population being divided concerning Q ≥ 0.5. Three subgroups, i.e., Q1, Q2 and Q3, included 34, 33 and 28 germplasms, respectively, and the remaining 10 germplasms were identified as the mixed population. During the experiment, 9 pairs of core primers were screened among the total of 12 for a complete differentiation regarding 105 tested materials, and the fingerprints of 91 C.×morifolium materials and 14 chrysanthemum-related materials were further constructed. Overall, there were significant genetic differences and rich genetic diversity among C.×morifolium materials, which would shed light on the garden application and variety selection fields of C.×morifolium. The fingerprint database of 105 C.×morifolium varieties and chrysanthemum-related species may provide technical support for future research regarding the identification and screening system of C.×morifolium varieties.