31种石斛属植物及金石斛遗传多样性SRAP分析及DNA指纹图谱研究

石斛属（Dendrobium Sw.）为兰科（Orchidaceae）兰亚科（Subfam. Orchidoideae）植物,在我国有着广泛的分布与应用,多用于名贵中药材和观赏植物,具有较高的经济价值。由于苛刻的生长条件加上过度的开发利用,导致野生石斛资源破坏严重,大部分种类已近枯竭,这也导致市场上出现的石斛种质混杂、真伪鉴定困难等问题亟待解决。石斛属植物遗传多样性的研究和数字指纹图谱的建立,可以为石斛的品种鉴定与分类、良种选择与保护提供理论依据,对保护药用植物生物多样性也具有十分重要的意义。以31种石斛属植物及金石斛（Flickingeria comata）为研究对象,应用SRAP-PCR分子标记技术对其开展遗传多样性分析并构建DNA指纹图谱。结果表明：利用筛选出来的15对引物进行扩增,共扩增出264个位点,其中多态性位点为251个,平均每对引物扩增出16.73个多态性位点,多态比率达95.08%,其中Me2-Em5、Me4-Em3、Me4-Em5、Me7-Em3、Me7-Ee7等5对引物的多态百分率均为100%,这些引物组合对石斛种类的鉴别效率较高;经过计算31种石斛属植物与金石斛间的观测等位基因数（N_a）为1.784,有效等位基因数（N_e）为1.430,Nei’s基因多样性指数（H）为0.202,Shannon’s信息指数（I）为0.386,石斛种间存在较高的遗传变异度与丰富的遗传多样性水平;其遗传相似系数的变化范围为0.591~0.851,亲缘关系较近,当遗传相似性系数为0.660时,金石斛单独聚为一类,当遗传相似性系数为0.724时,可将31个石斛属植物进一步分为10组;构建DNA指纹图谱可单独鉴别出31种石斛属植物以及金石斛,为石斛遗传背景分析和快速鉴别石斛种类提供科学依据。

SRAP Analysis and DNA Fingerprinting of Genetic Diversity of 31 Species of Dendrobium and Flickingeria comata

Dendrobium Sw. belongs to the Orchidaceae (Subfam. Orchidoideae), which has a wide distribution and application in China. Most are precious Chinese medicinal materials and ornamental plants with high economic values. Due to harsh growth conditions and excessive development and utilization, wild Dendrobium resources have been severely damaged, and most species have been nearly depleted, which also lead to problems such as mixed germplasm of Dendrobium in the market and difficulties in authenticity identification. The research on the genetic diversity of Dendrobium plants and the establishment of digital fingerprints can provide a theoretical basis for the identification and classification of Dendrobium species, the selection, and protection of fine varieties, and it is also of great significance to protect the biodiversity of Dendrobium resources. Taking 31 species of Dendrobium and Flickingeria comata as the research objects, the genetic diversity was analyzed by the SRAP-PCR molecular marker technology and DNA fingerprints were constructed. 15 pairs of primers were used for amplification, and a total of 264 loci were amplified, including 251 polymorphic loci, and an average of 16.73 polymorphic loci with a 95.08% ratio were amplified for each pair of primers. The polymorphism percentage of 5 pairs of primers including Me2-Em5, Me4-Em3, Me4-Em5, Me7-Em3, Me7-Ee7 was 100%, and the primer combinations had higher identification efficiency for Dendrobium species. After calculation, the number of observed alleles (N_a) between 31 species of Dendrobium and F. comata was 1.784, the number of effective alleles (N_e) was 1.430, the Nei’s gene diversity index (H) was 0.202, and Shannon’s information index (I) was 0.386, which showed there was a high degree of genetic variability and rich genetic diversity among Dendrobium species. The variation range of the genetic similarity coefficient was 0.591-0.851, and the genetic relationship was relatively close. When the genetic similarity coefficient was 0.66, Dendrobium could be clustered into one group. When the genetic similarity coefficient was 0.724, 31 Dendrobium species could be divided into 10 groups. The DNA fingerprints constructed by three pairs of primers, including Me1-Em3, Me3-Em3, and Me1-Em6, could identify 31 species of Dendrobium and F. comata independently, providing a scientific basis for the analysis of the genetic background and the rapid identification of D. species.