通过RAPD分析揭示五种淡水无齿蚌（软体动物门：双壳类）的遗传多样性

Unionidae(Bivalvia)are distributed infreshwaters,and represent a significanttaxonof benthic community[1].In China,freshwater mussels are abundant resources[2].Since1949,substantial investigations onthe unionidfau-na had been undertakenin China[3—8].Withreference tooverseas research[9,10],a preliminary reorganization onthe Unionidae was performed accordingtosome classifica-tion characteristics such as shell shape,larvae character-istics,and breeding habit[11].Due tothe serious conver-gence o…     

Genetic variability among freshwater mussel Anodonta woodiana (Lea, 1834) (Bivalvia: Unionidae) populations recently introduced in Poland

The alien Chinese mussel Anodonta woodiana was first reported in Poland in the system of heated lakes near Konin in 1993. Genetic studies with use of three molecular techniques (isoenzyme electrophoresis, PCR-RFLP and sequence analysis of a COI gene fragment) were carried out on the Polish first populations of A. woodiana. The studies have revealed low genetic variation between the populations (Nei's genetic distance for 12 loci ranged 0.000 to 0.007) as well as their considerable polymorphism. Each population averaged 2.28 alleles per locus, 2.72 alleles per polymorphic locus, and 75% polymorphic loci. Restriction analysis of the COI gene fragment have not revealed variability between the analysed specimens, including males and females. Restriction enzymes, ScrFI, Csp6I, and EcoRI used in the COI gene fragment PCR-RFLP generate distinct restriction patterns, which can be molecular markers for A. woodiana. The sequence obtained for COI fragment was the same in the examined female and male specimens and represents F mitotype (DNA was isolated from somatic tissues). The divergence between A. woodiana F and M mitotypes is high (34%), however it remains within the range of the general character of the DUI (doubly uniparental inheritance) phenomenon in freshwater bivalves (Unionidae).     

Isolation and characterization of microsatellite loci in western North American Anodonta species

We have developed and characterized 13 microsatellite loci from a group of Anodonta species in western North America, and demonstrated their utility in populations representing two major clades in this genus. Allelic diversity and polymorphic information content were high for all loci, although these characteristics varied across populations. Deviations from Hardy-Weinberg genotypic ratios were not detected, although the estimated frequency of null alleles was high in one population for one locus. This is the first set of microsatellite loci to be developed for freshwater mussels in western North America, and will be useful for describing gene flow patterns among populations.     

濒危植物三棱栎遗传多样性的RAPD分析

用随机扩增多态DNA (RAPD)标记对 5个三棱栎 (Trigonobalanusdoichangensis)居群共 99个个体进行遗传多样性和居群遗传结构分析。 16个引物共检测到 15 7个位点 ,其中多态位点 83个 ,占 5 2 87%。物种水平Shannon多样性指数I =0 2 4 31,Nei基因多样度h =0 15 95 ,种内总遗传变异量Ht=0 16 0 0 ,居群内遗传变异量Hs =0 0 74 9,居群间变异量大于居群内变异量 ,表明三棱栎的遗传变异主要存在于居群之间。与同科植物相比 ,三棱栎遗传多样性较低 ,遗传分化系数Gst =0 5 32 0 ,说明居群间的遗传变异占 5 3 2 0 % ,居群间已出现强烈的遗传分化。当地人的强烈活动造成的生境破碎化和居群隔离 ,以及三棱栎演化过程中的地史变化对其种群发展的影响等 ,可能是造成其居群间强烈的遗传分化和较低遗传多样性的原因。基于本研究结果 ,提出了三棱栎遗传多样性的保护策略。     

Extensive allozyme monomorphism in a threatened species of freshwater mussel, Margaritifera hembeli Conrad (Bivalvia: Margaritiferidae)

A threatened but under-studied component offreshwater biodiversity in North America is thenative freshwater mussels (Bivalvia:Unionoidea: Margaritiferidae and Unionidae). Genetic data suggest that these mussel speciesgenerally exhibit levels of variability similarto other invertebrates. We surveyed allozymevariation in the Louisiana Pearlshell, Margaritifera hembeli (Margaritiferidae), athreatened freshwater mussel. Five examinedpopulations are monomorphic for 25 allozymeloci, the first report of a native freshwatermussel species with extensive allozymemonomorphism. Low genetic diversity appears tobe characteristic of margaritiferids, as anANOVA indicated that mussels of the familyMargaritiferidae have significantly lowerlevels of heterozygosity than the mussels ofthe family Unionidae. Margaritiferids havestrong habitat preferences and modification ofhabitat leads to rapid loss of populations.Although bottlenecks are known to cause lowgenetic variability, margaritiferids mayexhibit meta-population structure withextinction/re-colonization dynamics leading tolow genetic variability. Margaritiferidsgenerally exhibit a patchy distribution with acolonization rate that is approximately twicethat of extinction. Tests of themetapopulation hypothesis will requireadditional allozyme population genetic data aswell as hypervariable microsatellite loci.     

濒危灌木长叶红砂遗传多样性的RAPD分析

应用RAPD分子标记对濒危灌木长叶红砂(Reaumuria trigyna)种群遗传多样性进行了分析.应用18条随机引物对长叶红砂5个种群的95个个体进行扩增,检测到118个位点,其中多态位点105个.结果表明:长叶红砂种群的多态位点比率(P)为88.98%,显示出长叶红砂种群存在较高的遗传多样性.Shannon多样性指数(0.4966)、Nei基因多样性指数(0.3303)和基因分化系数(Gst=0.1425)的分析结果显示,长叶红砂种群遗传变异大多存在于种群内,种群间的遗传分化占14.25%.聚类分析表明,长叶红砂种群遗传距离与地理距离之间无直接相关关系.遗传多样性水平与物种特性和所处不同群落有关,濒危植物并不一定表现为遗传变异水平的降低.     

Isolation and characterization of fifteen polymorphic microsatellite loci in the threatened freshwater mussel Solenaia oleivora (Bivalvia: Unionidae)

Freshwater mussels are among the most endangered fauna in the world, while little is known about their genetic diversity. We isolated 15 polymorphic microsatellite loci in a threatened freshwater mussel, Solenaia oleivora. The characteristics of these loci were tested with 46 individuals. The number of alleles per locus ranged from 2 to 15, and the PIC value ranged from 0.250 to 0.880. The observed and expected heterozygosity ranged from 0.167 to 0.929 and from 0.297 to 0.900, with an average of 0.562 and 0.647, respectively. Eight loci deviated significantly from Hardy–Weinberg equilibrium. These polymorphic microsatellites will be useful for the assessment of population genetic diversity and resource conservation of S. oleivora. Cross-species amplification demonstrated that six of 15 microsatellite loci amplified successfully in two other unionid species.     

三种摇蚊幼虫RAPD扩增条件的优化及在昆虫系统发育分析中的应用

为了研究摇蚊科昆虫种群遗传的多样性,以促进对其资源的合理保护,以萨摩亚摇蚊Chironomus samoensisEdwards基因组DNA为模板,对摇蚊幼虫的RAPD扩增条件进行优化,建立了摇蚊幼虫RAPD扩增反应的最佳体系:按照利用优化的RAPD扩增条件进行研究,实验有着良好的重现性。用16个随机引物对3种摇蚊幼虫类群各10个个体进行RAPD扩增,其中萨摩亚摇蚊共扩增出78个条带,多态座位率为41.03%,Shannon遗传多样性指数为0.2570,群体内相似度为0.8730;红裸须摇蚊Propsilocerus akamusi(Tokunaga)共75个条带,多态座位率为44.0%,Shannon遗传多样性指数为0.2472,群体内相似度为0.8731;刺铗长足摇蚊Tanypus punctipennis(Fabricius)共67个条带,多态座位率为41.79%,Shannon遗传多样性指数为0.1943,群体内相似度为0.9066。聚类分析结果表明,刺铗长足摇蚊与红裸须摇蚊的亲缘关系较近。     

Conservation genetics of endangeredBrasenia schreberi based on RAPD and AFLP markers

Brasenia schreberi J.F. Gmelin is a declared endangered species found in the lakes and ponds of South Korea. For planning its conservation strategy, we examined the genetic diversity within and among six populations, using randomly amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP). Polymorphisms were more frequently detected per loci with AFLP (69.3%) than RAPD (36.8%). High genetic diversity was recognized within populations: polymorphic loci (PPL) values ranged from 36.3% in the CJM population to 74.5% in the GGT population, with a mean value of 47.8% based on AFLP markers. Great genetic differentiation (θ^B) was detected among the six populations (0.670 on RAPD and 0.196 on AFLP), and we calculated a low rate of gene flow (N_em), i.e., 0.116 on RAPD and 0.977 on AFLP. Furthermore, a Mantel test revealed that no correlation existed between genetic distances and geographical distances among the six local populations, based on RAPD or AFLP markers. These results are attributed to a number of factors, including an insufficient length of time for genetic diversity to be reduced following a natural decline in population size and isolation, adaptation of the genetic system to small population conditions, and a restricted gene flow rate. Based on both its genetic diversity and population structure, we suggest that a strategy for conserving and restoringB. schreberi must focus on maintaining historical processes, such as high levels of outbreeding, while monitoring increased gene flow among populations. This is because a reduction in genetic diversity as a result of genetic drift is undesirable.     

Analysis of genetic diversity in red clover (Trifolium pratense L.) breeding populations as revealed by RAPD genetic markers.

Red clover is an important forage legume species for temperate regions and very little is known about the genetic organization of its breeding populations. We used random amplified polymorphic DNA (RAPD) genetic markers to address the genetic diversity and the distribution of variation in 20 breeding populations and cultivars from Chile, Argentina, Uruguay, and Switzerland. Genetic distances were calculated for all possible pairwise combinations. A high level of polymorphism was found and the proportion of polymorphic loci across populations was 74.2%. A population derived from a non-certified seedlot displayed a higher proportion of polymorphic loci than its respective certified seedlot. Gene diversity values and population genetics parameters suggest that the populations analyzed are diverse. An analysis of molecular variance (AMOVA) revealed that the largest proportion of variation (80.4%) resides at the within population level. RAPD markers are a useful tool for red clover breeding programs. A dendrogram based on genetic distances divided the breeding populations analyzed into three distinct groups. The amount and partition of diversity observed can be of value in identifying the populations that parents of synthetic cultivars are derived from and to exploit the variation available in the populations analyzed.     

大猿叶虫四地理种群遗传多样性的RAPD分析

以大猿叶虫Colaphellus bowringi Baly 4个地理种群的基因组DNA为材料, 进行RAPD分析。从80条引物中筛选出11条稳定性好、多态性高的引物进行扩增, 共得到65个扩增位点, 53个多态位点, Nei氏遗传多样性指数为0.1049~0.2061, Shannon多样性指数为0.1641~0.3167。结果表明所分析的大猿叶虫遗传变异很高, 其中江西龙南种群遗传变异最小, 山东泰安种群遗传变异最高。种群间的遗传距离范围为0.0636~0.3200, 其中江西龙南种群和江西修水种群间的遗传距离最小, 哈尔滨种群与江西龙南种群间的遗传距离最大, 种群遗传距离的大小与其相对地理距离的远近吻合。结果提示种群遗传距离的大小与它们生物学上的相似性有关联。     

濒危植物资源冷杉遗传多样性研究

资源冷杉(Abies ziyuanensis)属松科(Pinaceae)冷杉属(Abies),是国家一级保护的濒危植物。利用RAPD分子标记对分布于广西资源县银竹老山和湖南省炎陵县大院的2个资源冷杉种群共54株进行了遗传多样性分析。用20个随机引物共扩增到126个位点,其中66个位点是多态性的,总的多态位点百分率为52.4％,在两个种群分别为33.6％和31。2％。Nei's基因多样性为0.312,Shannon信息指数为0.475,两个种群的遗传分化系数Gst为0.455。基于Nei's遗传距离进行了UPGMA聚类分析,结果两种群的样品彼此明显区分开来。分析结果表明资源冷杉种群内的遗传多样性较低,但两种群间的遗传分化明显。需要同时重视两分布地的资源冷杉的保护工作。     

Population Genetic Structure of Astyanax scabripinnis (Teleostei, Characidae) from an Urban Stream

Despite the great anthropogenic interference on urban streams, information is still scarce about the genetic variability and structure of native fish populations inhabiting such streams. In the present study, random amplified polymorphic DNA (RAPD) markers were used to analyze genetic variability and structure of populations assigned to the Neotropical fish species Astyanax scabripinnis from an urban stream located in Londrina, Paraná State, southern Brazil. Thirty individuals of this species were collected from three sites throughout the upper Cambé stream. A total of 10 primers amplified 159 loci, of which 128 (80.5%) were polymorphic. Each of the three populations showed very similar proportions of polymorphic loci, which ranged from 63.5 to 64.8%. Unbiased genetic distances varied from 0.0612 to 0.0646. Theta_p-test values indicated moderate to high genetic differentiation among individuals from different localities. The number of migrants varied from 1.34 to 1.46, suggesting a low gene flow between populations. The genetic similarity among all individuals studied ranged from 0.424 to 0.848. The results suggest that populations of A. scabripinnis in Cambé stream are undergoing genetic differentiation.     

河蚌血细胞对细菌趋化移动的初步研究

采用改进的毛细管法 ,研究了圆背角无齿蚌 (Anodontawoodianapacifica)和三角帆蚌 (Hyriopsiscum ingii)两种淡水河蚌离体血细胞对两种水体中常见病原细菌的趋化移动作用 ,及血清对其的影响。结果显示 ,两种河蚌的离体血细胞对细菌都具有趋化移动作用 ,产生趋化移动的血细胞数量都显著高于无细菌的对照组 (P <0 0 5 )。在有血清时 ,血细胞对荧光极毛杆菌 (Pseudomonasfluorescens)的趋化移动活性略高于肠型点状气单孢菌 (Aeromonaspunctataf.intestinalis) ,圆背角无齿蚌离体血细胞的趋化移动能力显著高于三角帆蚌 (P <0 0 5 )。血清对河蚌离体血细胞的趋化移动作用有显著的促进作用 (P <0 0 5 )。     

Development of twelve polymorphic microsatellite loci in polyploid endangered Omphalogramma vincaeflora Franch. (Primulaceae)

A total of twelve polymorphic microsatellite loci were developed from polyploid endangered species, Omphalogramma vincaeflora (Primulaceae). These loci were screened for variability among 45 individuals from three populations in China. The primers amplified loci with allele number ranging from 3 to 9, with an average of 4.25 per locus. Polymorphism information content ranged from 0.23 to 0.86. Nei’s genetic diversity ranged from 0.34 to 0.86. These primers provide an opportunity to use polymorphic DNA markers to study the population genetic structure and its breeding system in this species.     

广东省十种卷柏属植物RAPD初步分析

采用RAPD标记对卷柏属10种植物进行了分类鉴定、种间亲缘关系及遗传多样性的初步分析。结果显示:10个RAPD引物很好地区分了10个种,每个种均有2~12个种单态特征位点;UPGMA聚类树与有关文献表型分类结果不尽一致;卷柏属不同种具有不同的遗传多样性水平。     

新疆吐鲁番荒漠土壤绿藻的多样性分析

应用RAPD技术对吐鲁番地区火焰山及艾丁湖区域分离的15株土壤绿藻(chlorophyta)品系的遗传多样性及其亲缘关系进行探讨。结果表明:从20个随机引物中,筛选出多态性和重复性较好且谱带清晰的引物8个,这8个引物扩增出的DNA片段大多在300～2 000 bp之间,所形成的多态性位点数差距较大,显示该区域土壤绿藻具有较丰富的遗传多样性;15株土壤绿藻扩增共得到74条谱带,71条多态性带,其多态性比率为95.95%;聚类分析显示15株土壤绿藻明显地聚为2大类,与其来源相对应,即隶属于同一亚组或相近亚组的不同种基本归为一类,其种间关系与传统的形态学分类结果相吻合。     

地被植物鹅绒委陵菜的遗传多样性研究

采用RAPD方法对不同海拔和植被盖度下的鹅绒委陵菜（Potentilla anserinaL.）6个种群的遗传多样性水平进行了研究。结果表明,13条随机引物共扩增出132条带,多态性位点为117;鹅绒委陵菜物种水平的遗传多样性较高,且种群间分化明,多态位点比率（88.64%）、Nei’s基因多样性（0.3180）、Shannon信息多样性指数（0.4732）均远高于多数克隆植物。6个种群内的遗传多样性水平变化较大,多态位点在各种群中分布不均衡,种群间分化系数（GST）为47.7%,基因流较低（Nm=0.5482）,多态位点比率在31.06%～74.24%之间,Shannon信息指数在0.1711～0.3625之间,Nei基因多样性指数在0.1164～0.2425之间。多样性水平的变化与海拔没有明显的相关性,而与生境盖度呈显著正相关。从而推断在盖度低、资源丰富的环境中,该物种可能更倾向于克隆繁殖。     

Genetic variation and population structure of endemic yellow catfish, <Emphasis Type="Italic">Horabagrus</Emphasis><Emphasis Type="Italic">brachysoma</Emphasis> (Bagridae) among three populations of Western Ghat region using RAPD and microsatellite markers

Random amplified polymorphic DNA (RAPD) and microsatellite markers were applied to evaluate the genetic variation in endemic and endangered yellow catfish, Horabagrus brachysoma sampled from three geographic locations of Western Ghat, South India river systems. In RAPD, of 32 10-mer RAPD primers screened initially, 10 were chosen and used in a comparative analysis of H. brachysoma collected from Meenachil, Chalakkudy and Nethravathi River systems. Of the 124 total RAPD fragments amplified, 49 (39.51%) were found to be shared by individuals of all 3 populations. The remaining 75 fragments were found to be polymorphic (60.48%). In microsatellites, six polymorphic microsatellite loci were identified by using primers developed for Pangasius hypophthalmus, Clarias macrocephalus and Clarias gariepinus. The identified loci were confirmed as microsatellite by sequencing after making a clone. The nucleotide sequences of 6 loci were published in NCBI genbank. The number of alleles across the six loci ranged from 4 to 7 and heterozygosities ranged from 0.07 to 0.93. The mean number of alleles and effective number of alleles per locus were 5.00 and 3.314, respectively. The average heterozygosity across all investigated samples was 0.72, indicating a significant deficiency of heterozygotes in this species. RAPD and microsatellite methods reported a high degree of gene diversity and genetic distances depicted by UPGMA dendrograms among the populations of H. brachysoma.     

凉水国家自然保护区天然红松林遗传变异的RAPD分析

采用随机扩增多态DNA(random amplified polymorphic DNA RAPD)技术,研究了凉水国家自然保护区天然红松林的遗传变异水平及其分布规律。16个10bp长度的随机引物在8个实验样地内72个个体中共检测了96个位点,其中55个是多态位点。在种水平上,红松的多态位点比率为P=58.51%;Nei's遗传变异指数为H=0.2868;Shannon's指数为I=0.3654,所有的变异中,有92.47%的变异存在于样地内, 7.53%的变异存在于样地间。根据各遗传多样性在不同样地的分布,初步探讨了红松遗传变异水平与其种群发生及生境的相互关系。