佛坪国家级自然保护区秦岭箭竹克隆结构的SSR分析

Clonal structure of a Fargesia qinlingensis population inferred using simple sequence repeat fingerprints in Foping National Nature Reserve

利用SSR分子标记技术分析了佛坪国家级自然保护区秦岭箭竹(Fargesia qinlingensis)的克隆多样性和克隆结构,以探讨小尺度范围内秦岭箭竹自然居群遗传变异的分布特征,对该种开花特性、高山地区生态环境维护和大熊猫的保护提供重要依据。结果表明7对SSR引物共扩增出79个位点,其中多态性位点77个,多态位点百分率(PPB)为97.47%。秦岭箭竹的142个分株共形成107个克隆,最大克隆可达5 m。克隆多样性略高于其他克隆植物的平均值(D=0.62,G/N=0.17,E=0.68),基因型比率(G/N)、Simpson指数(D)、平均克隆大小(N/G)和Fager均匀性指数(E)分别为0.7535、0.9680、1.3271和0.5109。克隆空间结构分析表明秦岭箭竹的克隆构型为密集型,各克隆呈镶嵌性分布,同一克隆的分株排列紧密。克隆聚类分析表明各克隆之间聚类不明显,总体上来自同一样地的克隆被聚为一类。空间自相关分析显示在空间距离为36 m范围内,分株比基株有更显著的空间遗传结构,空间自相关系数r的取值范围分别为0.084—0.626和0.024—0.288,说明克隆繁殖在一定程度上限制了空间遗传结构的范围。样地内秦岭箭竹个体在空间距离小于44 m时存在显著的正相关空间结构,特别是在4 m处表现出最大的空间自相关系数(r=0.626),表明空间距离相距4 m内的个体最有可能属于同一克隆,4 m比5 m更能表现出清晰的克隆结构,X-轴截距为52.280,代表了秦岭箭竹不规则克隆的平均最小长度。秦岭箭竹的克隆多样性和克隆结构与初始苗补充、花粉散播方式和微环境差异有关。

The clonal diversity and clonal structure of a Fargesia qinlingensis population densely distributed in Foping National Nature Reserve were analyzed by simple sequence repeat （SSR） fingerprints. We aimed to describe how the clonal structure of F. qinlingensis was established at a small scale and its association with flowering traits, ecological environment conservation in high mountains, and the protection of giant pandas. In all, 81 SSR primers were designed, of which 7 pairs with good stability, high polymorphism, and specificity for our research were selected for the 142 sampled ramets. These 7 SSR primers generated a total of 79 valid loci, of which 77 （97.47%） were polymorphic. We successfully genotyped 107 clones among the 142 sampled ramets. The largest single clone may cover a spatial distance of approximately 5 m. This species showed a high clonal diversity, with the proportion of distinguishable genotypes （G/N）, Simpson＇s index of diversity （D）, average size of genotypes （N/G） , and Fager＇s evenness （E） determined as 0.7535, 0.9680, 1.3271, and 0.5109, respectively. Its clonal diversity was slightly higher than the average diversity of other clonal plants, with a mean Simpson＇s index of diversity （D） = 0.62, proportion of distinguishable genotypes （G/N） = 0.17, and Fager＇s evenness （E） = 0.68. Analysis of clonal structure demonstrated that the spatial distribution pattern of F. qinlingensis exhibited a phalanx growth strategy at the ramet level, in contrast with our results that showed ramets belonging to the same clone were closely aggregated and formed distinct clumps at the 1 m × 1 m sample scale and clones were juxtaposed at the 5 m × 5 m sample scale. Although unweighted pair-group method analysis （UPGMA） demonstrated no distinct clusters of clones, the clones in the same plot were always classified into the same clade. Spatial autocorrelation analysis showed the spatial autocorrelation coefficientsr were 0.084-0.626 and 0.024-0.288 at the ramet and genet levels, respectively. This result indicated a significantly stronger spatial autocorrelation at the ramet level than at the genet level for F. qinlingensis within a spatial distance of 36 m, which implied that in spite of pollen flow might extend the spatial genetic structure, clonal propagation made a certain restriction of the spatial genetic structure. The autocorrelation coefficient was significantly positive within the distance of 44 m at the ramet level and the X-intercept representing the average minimum length of irregular clone was 52.280 m. In addition, the largest spatial autocorrelation coefficient was 0.626 at 4 m, which suggested that ramets within 4 m most likely belonged to the same clone. It also implies that a sampling scale of 4 m shows a more distinct clonal structure than that of 5 m. Furthermore, we detected significant negative autocorrelation from 56 m to 116 m, but no significantr-values were detected beyond 120 m. At the genet level, the autocorrelation coefficient was significantly positive within the distance of 36 m, significantly negative from 56 m to 104 m, but showed no significant autocorrelation from 36 m to 56 m, and beyond 108 m. Our results revealed that clonal diversity and clonal structure could be affected by initial seeding recruitment, pollen dispersal, and heterogeneity of microenvironment. A relatively larger sample size and a more reasonable sample strategy would be favorable to investigate a more distinct clonal structure and clonal diversity of F. qinlingensis in different habitats.