DNA标记技术在螨类系统学研究中的应用

长期以来,螨类主要依靠其形态特征进行系统学研究。DNA标记是指能反映生物个体或物种间基因组中某种差异特征的DNA片段。近年来,DNA标记技术在螨类系统学研究中得到越来越广泛的应用。本文综述了随机扩增多态性RAPD、限制性内切酶片段长度多态性RFLP、微卫星SSR、核酸序列扩增、扩增片段长度多态性AFLP和直接扩增片段长度多态性DALP等6种DNA标记技术在螨类系统学研究中的应用现状及前景。     

云南丽江山慈菇遗传多样性的DALP分析

采用DALP (Direct amplification of length polymorphism) 分子标记技术, 对产自云南的药用植物丽江山慈菇Iphigenia indica (L.) Kunth的9个居群进行DNA指纹检测。筛选出5个引物组合, 扩增共产生131条DNA片段, 其中104 条谱带具有遗传多态性, 约占79 39%, 平均每组引物扩增所得多态条带为20 8, 9个居群平均多态百分率为42 21%。9个居群平均观察等位基因数Na为1 4224, 总Na为1 7939; 平均有效等位基因数Ne 为1 3141, 总Ne 为1 4810; 平均遗传多样性指数H为0 1745, 总H为0 2831; 平均Shannon 多样性指数I 为0 2527, 总I为0 4231; 总基因多样性Ht为0 2831, 居群内多样性Hs 为0 1745, 居群间基因分化系数Gst为0 3834, 即丽江山慈菇有61 66%的遗传变异来自居群内, 38 34%来自居群间, 居群间存在较高水平的遗传分化。滇西北居群的遗传多样性明显高于滇中居群的遗传多样性, 这与滇中地区丽江山慈菇野生资源被大规模挖掘有着直接的关系。     

A Study of Genetic Structure of Stephania yunnanensis (Menispermaceae) by DALP

Genetic diversity and genetic structure within and among ten populations of Stephania yunnanensis H. S. Lo and three populations of S. epigaea H. S. Lo from Yunnan province were evaluated by direct amplification of length polymorphism (DALP) markers. Five primer groups were screened, and a total of 287 DNA fragments were amplified, among which 266 were polymorphic, averaging 53.2 polymorphic bands per primer group in S. yunnanensis. The percentage of polymorphic bands of S. yunnanensis was 92.68% at the species level and 61.92% within the ten populations sampled. At the species level, the observed number of alleles (N _a) was 1.9268 and the effective number of alleles (N _e) was 1.5933; Nei’s gene diversity (H) was 0.3414; Shannon’s information index (I) was 0.5057. At the population level, N _a = 1.6192, N _e = 1.4001, H = 0.2298, and I = 0.3401. Total gene diversity of S. yunnanensis was 0.3419. Gene diversity within population was 0.2298, coefficient of gene differentiation was 0.3278, and estimated gene flow was 1.0254. The results indicated that the genetic differentiation was relatively higher among populations of S. yunnanensis. DALP markers were an informative and useful method for assaying genetic diversity and authenticating species of Stephania. These data could provide basic molecular evidence for establishing a reasonable strategy for protecting and exploiting the resource of S. yunnanensis.     

汤洪敏云南野生大白口蘑遗传多样性研究

应用DALP技术对云南野生大白口蘑遗传多样性进行分析。筛选的5组引物在大白口蘑6个菌塘的56朵子实体中检测到202个位点,其中多态位点151个,多态位点百分比PPB为74.75%。6个菌塘总的观察等位基因数Na为1.7475;总的有效等位基因数Ne为1.4659;Nei’s基因多样性指数H为0.2722;总的Shannon多样性指数I为0.4053;菌塘总的基因多样度Ht为0.2705,菌塘内基因多样性度Hs为0.012,菌塘间基因分化系数Gst为0.9556,菌塘的遗传一致度在0.60~0.80之间。结果表明大白口蘑遗传变异有95.56%存在菌塘间,4.44%存在菌塘内,即大白口蘑遗传变异绝大多数存在于菌塘之间。     

Genetic diversity and population structure of the endangered species Psammosilene tunicoides revealed by DALP analysis

Psammosilene tunicoides W. C. Wu et C.Y. Wu (Caryophyllaceae) is a rare plant endemic to China. We used the direct amplification of length polymorphism (DALP) method to analyse the genetic diversity of 17 natural populations of this species from Yunnan, Sichuan and Guizhou provinces. The percentage of polymorphic bands (PPB) was 99.1% at the species level and 19.9% at the population level. The total genetic diversity (Ht) was 0.2827; the genetic diversity within populations (H_s) was 0.0594 and the coefficient of gene differentiation (G_st) was 0.7899. The results indicated low levels of genetic diversity within populations and extremely high genetic differentiation among populations. This pattern might be caused by genetic drift resulting from intensive harvesting and habitat fragmentation. It might also be a result of local adaptation. We further performed experiments on its breeding system and found a high rate of selfing in P. tunicoides, which may be partially responsible for its population structure.     

莲瓣兰DALP遗传多样性研究

采用DALP(Direct amplification of length polymorphism)检测莲瓣兰5个居群的遗传结构.5个引物组合共检测到103个多态位点.和其它具有相似生活史(多年生草本、以动物为媒介的杂交、种子随风散布)的物种相比,莲瓣兰原变种水平(PPB=88.18％,A=1.8818,Ae=1.4880,H=0.2911,I=0.4412)和物种水平(PPB=93.64％,A=1.9364,Ae=1.5262,H=0.3129,I=0.4732)均具有较高的遗传多样性;各居群间(Gst=0.3292)存在较大的遗传分化.基因流的估计值(Nm=1.0186)表明莲瓣兰物种水平上各居群间每代有1.0186个移居个体.地理隔离和选择压力可能是造成莲瓣兰居群间基因流动受到限制的原因.在这些研究的基础上,探讨了野生莲瓣兰资源的保护问题.     

云南野生大白口蘑遗传多样性研究

应用DALP技术对云南野生大白口蘑遗传多样性进行分析。筛选的5组引物在大白口蘑6个菌塘的56朵子实体中检测到202个位点,其中多态位点151个,多态位点百分比PPB为74.75%。6个菌塘总的观察等位基因数Na为1.7475;总的有效等位基因数Ne为1.4659;Nei’s基因多样性指数H为0.2722;总的Shannon多样性指数I为0.4053;菌塘总的基因多样度Ht为0.2705,菌塘内基因多样性度Hs为0.012,菌塘间基因分化系数Gst为0.9556,菌塘的遗传一致度在0.60~0.80之间。结果表明大白口蘑遗传变异有95.56%存在菌塘间,4.44%存在菌塘内,即大白口蘑遗传变异绝大多数存在于菌塘之间。     

Genetic differentiation within metapopulations of Euphydryas aurinia and Melitaea phoebe in China

We analyzed genetic differentiation within metapopulations of two species of checkerspot butterfly, Euphydryas aurinia and Melitaea phoebe, in China. To generate genetic information, we used a new molecular technique, DALP – direct amplified length polymorphism. AMOVA results showed that most of the variation occurred among individuals within local populations of both E. aurinia and M. phoebe. However, while there was differentiation among local population in E. aurinia (P < 0.001), there was no subdivision in metapopulation of M. phoebe (P = 0.210). This is consistent with the behavior of M. phoebe adults being more dispersive than E. aurinia. Within the M. phoebe metapopulation, three neighboring patches were always occupied during the observation period (1998–2000). In addition, the number of individuals in these three populations accounted for the majority of M. phoebe larvae, and hence we conclude that the M. phoebe metapopulation might exist as a source-sink metapopulation. On the other hand, the E. aurinia metapopulation is an example of a classical metapopulation. Therefore, the conservation management of these two species should reflect these differences.     

螨类系统学研究中的分子标记

近年来,分子标记技术在螨类系统学研究中起到越来越重要的作用。文章就几种螨类系统学中用到的分子标记的原理和应用作一回顾,其中包括随机扩增多态性RAPD、限制性内切酶片段长度多态性RFLP、直接扩增片段长度多态性DALP、扩增片段长度多态性AFLP、微卫星DNASSR、核酸序列分析。讨论这几种分子标记在其应用中的优势及其局限性,并对分子标记在螨类系统学研究中的应用作出展望。