基于SSR标记探讨三种金花茶植物的遗传多样性和遗传结构

薄叶金花茶、小花金花茶和小瓣金花茶是三种濒危金花茶植物,为了解珍稀濒危植物遗传多样性和遗传结构,该研究利用微卫星标记对他们的7个种群共184个个体进行了遗传多样性和遗传结构分析。结果表明:11个位点共检测到等位基因92个。在物种水平上,小瓣金花茶平均等位基因数(N_A)为3.9、有效等位基因数(N_E)为2.328、观测杂合度(H_o)为0.520、期望杂合度(H_e)为0.501,高于薄叶金花茶和小花金花茶。在种群水平上,有效等位基因数(N_E)在1.788～2.466之间,期望杂合度(H_e)在0.379～0.543之间;种群间遗传分化系数(FST)在0.143 7～0.453 3之间,种群间基因流(N_m)在0.301 5～1.488 9之间。AMOVA分子变异分析显示65.72%的变异存在于种群内。三种金花茶具有较低水平的遗传多样性和高水平的种群间遗传分化。STRUCTURE和PCoA种群遗传结构分析结果将取样种群分为2组,即薄叶金花茶和小花金花茶大部分个体分为一组,小瓣金花茶大部分个体分为一组。现存所有种群应根据实际情况尽快采取就地保护或迁地保护措施。     

板栗和锥栗同域居群的空间遗传结构

采用微卫星技术研究了板栗(Castanea mollissima)和锥栗(C. henryi)同域分布居群的空间遗传结构, 探讨了它们的遗传变异空间分布特征及其形成机制。结果表明,采用的7个微卫星在两物种中共扩增出173个等位基因,两物种都具有较高的遗传多样性且种间遗传分化水平较低(F_ST=0.051), 但空间自相关分析揭示板栗和锥栗在同域居群中具有不同的空间遗传结构,锥栗在100 m内有空间遗传结构,而板栗没有;同时,基于亲缘关系系数F_ij的Sp统计值也显示锥栗具有比板栗更强的空间遗传结构(板栗的Sp=0.002;锥栗的Sp=0.018)。居群遗传变异的空间结构是其传粉和种子散播及生境共同作用的结果,其中种子的近距离散播和居群密度可能是主要的因素。板栗和锥栗居群在小尺度上空间遗传结构的差异可能反映了它们种子的大小及扩散过程的差异。     

Phylogeny of Castanea (Fagaceae) based on chloroplast trnT-L-F sequence data

Species in the genus Castanea are widely distributed in the deciduous forests of the Northern Hemisphere from Asia to Europe and North America. They show floristic similarity but differences in chestnut blight resistance especially among eastern Asian and eastern North American species. Phylogenetic analyses were conducted in this study using sequences of three chloroplast noncoding trnT-L-F regions. The trnT-L region was found to be the most variable and informative region. The highest proportion of parsimony informative sites, more and larger indels, and higher pairwise distances between taxa were obtained at trnT-L than at the other two regions. The high A+T values (74.5%) in the Castanea trnT-L region may explain the high proportion of transversions found in this region where as comparatively lower A+T values were found in the trnL intron (68.35%) and trnL-F spacer (70.07%) with relatively balanced numbers of transitions and transversions. The genus Castanea is supported as a monophyletic clade, while the section Eucastanon is paraphyletic. C. crenata is the most basal clade and sister to the remainder of the genus. The three Chinese species of Castanea are supported as a single monophyletic clade, whose sister group contains the North American and European species. There is consistent but weak support for a sister–group relationship between the North American species and European species.     

Genetic diversity and population differentiation of chestnut blight fungus, Cryphonectria parasitica, in China as revealed by RAPD

Seventeen Cryphonectria parasitica populations sampled from six regions in China were investigated using RAPD. Across all 169 isolates from the 17 populations evaluated, 52 of the 71 markers (73%) were polymorphic, total genetic diversity (h) was 0.1463, and Shannon’s index was 0.2312. Diversity within populations accounted for 74% of total genetic diversity, and genetic differentiation among populations was 0.26 (G _ST = 0.26). Gene flow was 1.4 among the populations; higher gene flow was found among populations within regions and among regions [N _m (G _SR) = 2.8 and N _m (G _RT) = 3.5]. The unweighted pair group mean analysis (UPGMA) dendrogram revealed two distinct clusters: the northern China group and the southern China group. The spatial autocorrelation analysis revealed that the variation at most loci was randomly distributed and lacked spatial structure, but several loci and closer distances were spatially structured. Human activity and habitat could also be important factors affecting genetic structure among C. parasitica populations in China. Genetic diversity was highest in Southwest China, descending in an orderly fashion to Northeast China. This pattern indicated that Southwest China might be the center of origin of C. parasitica in China. The present study provides useful information for understanding the origin and spread of chestnut blight fungus in China and valuable data for formulating relevant strategies for controlling the disease in China.     

Allozyme investigations on the genetic differentiation between closely related pines —Pinus sylvestris, P. mugo, P. uncinata, andP. uliginosa (Pinaceae)

In eight natural European populations of four closely related taxa of pines (Pinus sylvestris, P. mugo, P. uncinata, andP. uliginosa) starch-gel electrophoreses revealed altogether 58 alleles at 15 loci from nine enzyme systems. With Nei's genetic distance (D) the largest mean genetic distance (D = 0.171) was found betweenP. sylvestris andP. mugo, a distance corresponding to that between other closely related pine species. Mean genetic distances between the other taxa were less than half that value and characteristic for subspecies or varieties rather than for species. On the basis of our results we suggest that both,P. uliginosa andP. uncinata, could be the result of the ancient hybridization betweenP. sylvestris andP. mugo.     

Population structure,gene diversity,and differentiation in natural populations of Cedar pines (Pinus subsect.Cembrae,Pinaceae) in the USSR

Twelve natural populations of four cedar pine species,Pinus sibirica, P. cembra, P. pumila, andP. koraiensis, occurring in the Soviet Union were investigated by starch-gel electrophoresis. Frequencies of 55 alleles at 19 loci were determined. Interpopulation genetic diversity inP. sibirica andP. pumila was only 2–4 per cent of the total genetic diversity. Nei's distance coefficient (Dn) was used to estimate the level of genetic differentiation among conspecific populations and among species. Dn values among populations ranged from 0.006 to 0.038. A dendrogram constructed using Dn values divided cedar pines species into 2 clusters:sibirica-cembra (Dn = 0.030) andpumila-koraiensis (Dn = 0.143). Nei's distance between these clusters was 0.232. On the basis of the data obtained it was possible to draw the following conclusion:P. sibirica, P. pumila, andP. koraiensis are distinct species, whileP. cembra should apparently be regarded as geographicalP. sibirica race.     

Genetic variation and phylogeographic history of <Emphasis Type="Italic">Picea likiangensis</Emphasis> revealed by RAPD markers

Repeated cycles of retreat and recolonization during the Quaternary ice ages are thought to have greatly influenced current species distributions and their genetic diversity. It remains unclear how this climatic oscillation has affected the distribution of genetic diversity between populations of wind-pollinated conifers in the Qinghai-Tibetan region. In this study, we investigated the within-species genetic diversity and phylogenetic relationships of Picea likiangensis, a dominant forest species in this region using polymorphic DNA (RAPD) markers. Our results suggest that this species has high overall genetic diversity, with 85.42% of loci being polymorphic and an average expected heterozygosity (H _E) of 0.239. However, there were relatively low levels of polymorphism at population levels and the differences between populations were not significant, with percentages of polymorphic bands (PPB) ranging from 46.88 to 69.76%, Nei’s gene diversity (H _E) from 0.179 to 0.289 and Shannon’s indices (Hpop) from 0.267 to 0.421. In accordance with our proposed hypothesis, a high level of genetic differentiation among populations was detected based on Nei’s genetic diversity (G _ST = 0.256) and AMOVA analysis (Phi _st = 0.236). Gene flow between populations was found to be limited (Nm = 1.4532) and far lower than reported for other conifer species with wide distribution ranges from other regions. No clusters corresponding to three morphological varieties found in the south, north and west, respectively, were detected in either UPGMA or PCO analyses. Our results suggest that this species may have had different refugia during the glacial stages in the southern region and that the northern variety may have multiple origins from these different refugia.     

Development of Novel Microsatellite DNA Markers from the Pacific Oyster Crassostrea gigas

We document the potential of novel microsatellites as a genetic tool in furthering our understanding of the Crassostrea gigas genetic structure. From the microsatellite-enriched libraries we constructed, 123 repeat regions that had sufficient sequence information to design polymerase chain reaction primer sets were isolated. From these, 9 primer pairs were screened in a C. gigas population of 67 individuals to evaluate the genetic variability. All but 1 of the 9 loci showed allelic variation (number of alleles, 2–20; observed heterozygosity, 0.119–0.925; unbiased expected heterozygosity, 0.139–0.914). Considerable discrepancy of genotypic proportions from the Hardy-Weinberg equilibrium was observed at 1 locus with an apparent heterozygote deficiency. Several loci were successfully amplified in 3 other related species with the appropriate allele size: 6 loci in C. sikamea, 4 loci in C. ariakensis, and 5 loci in C. nippona.     

基于SSR标记的大明松天然群体遗传多样性分析

大明松是广西和贵州特有的高山松类,具有很高的经济价值和生态价值,其自然种群长期没有得到充分的保护和利用,不利于该树种长期稳定发展。为了合理保护和开发利用大明松天然遗传资源,该文利用12个SSR分子标记对大明松3个天然群体进行遗传多样性研究,分析其群体间的遗传分化和基因流,为该物种的保护策略提供参考。研究结果表明:(1)12对引物共检测到37 个等位基因,多态位点百分率为100%。每个位点平均观察等位基因数(Na)为3.08,平均有效等位基因数(Ne)为1.68, 不同位点间有效等位基因数差异较大; 每个位点平均观察杂合度(Ho)为0.35,平均期望杂合度(He)为0.40,平均多态信息含量(PIC)为0.31。(2)3 个群体的Shannon''s多样性指数变化范围为0.48～0.65,Nei''s基因多样度的变化范围为0.27～0.39,与其他近缘种松类相比遗传多样性偏低。群体平均观察杂合度为0.40,平均期望杂合度为0.33,平均有效等位基因数为1.58。群体间基因分化系数(Gst)为0.10,群体间的遗传分化水平较低,大部分变异均存在群体内。群体间的基因流(Nm)为2.74,说明大明松群体间的基因交流比较充分。该研究可为大明松生物多样性保护提供重要参考依据,为科学利用大明松资源打下基础。     

Allozyme variation in the diploid (A genome) populations ofScilla scilloides (Hyacinthaceae)

Allozyme variation at eleven loci encoding seven enzyme systems were examined in 20 populations of diploid (genome AA, 2n = 16)Scilla scilloides in China. In comparison with the average species of seed plants studied, populations of this species display a high amount of genetic variation (A = 2.0, P = 58.6%, H_o = 0.172, and H_e = 0.185). Allozyme variation pattern revealed predominant outcrossing within populations and considerable differentiation (F_ST = 0.314) among populations as well as between the subtropic and temperate regions. The wide distribution, long existence and outcrossing are presumably the main factors responsible for the high genetic diversity within populations. But the gravity dispersal of seeds and pollination by small insects set limits to the increase of genetic variation within populations and promote differentiation between populations and regions. In addition, allozyme variation does not distinguishS. scilloides var.albo-viridis and suggests that subtropic populations may be considered as a genetic entity.     

基于SSR分子标记的延胡索遗传多样性研究

采用SSR分子标记分析延胡索的遗传多样性,筛选出多态性高、稳定性好的12对引物,对19个居群360份延胡索样品进行了群体遗传分析。结果表明:(1)12对SSR引物共扩增出多态性位点227个,检测到4~9个等位基因,平均等位基因数目为5.25个,表现出丰富的多态性;延胡索居群具有较高的遗传多样性(Na=5.25,I=1.192 6,H=0.387 9),物种间遗传分化程度高(Fst=0.388 3),基因流较弱(Nm=0.393 8)。(2)UPGMA聚类分析和贝叶斯距离分析结果表明,19个居群明显聚为4大支;Mantle Test结果(r=0.326,P=0.01)表明,地理位置相近的种群亲缘关系更近。研究认为,延胡索的遗传结构是该物种自交为主的繁育系统、克隆生长、地理隔离以及居群间有限的基因流共同作用的结果,故对该物种的保护应以就地保护为主。     

Protein polymorphism inEligmodontia typus. Genetic divergence with other phyllotine cricetids

By means of gel electrophoresis of tissue extracts we have studied protein polymorphism inEligmodontia typus. The analysis was performed on specimens from five population samples collected at different sites in Patagonia (Argentina). Mean heterozygosity (\-h) and proportion of polymorphic loci (P) were determined on the basis of 19 loci. Considering all individuals as one sample, \-h gave a value of 0.16 and P was 70%. Although these values are much higher than those reported for most rodent species, they are very similar to those obtained by us for four species of the genusCalomys and forGraomys griseoflavus. There is a striking genetic identity (I_N=0.99) among populations from regions with different environmental conditions, indicating that the species possesses a common genic pool. Genetic distance with other species of the Phyllotini was estimated. D_N was lower betweenE. typus andCalomys (mean D_N=0.88) than betweenE. typus andGraomys griseoflavus (D_N=1.01). The high morphological similarity between these last two species, especially regarding those characters related to desert life adaptation, could be assigned, at least in part, to convergent evolution.     

Allozyme Variation and Phylogenetic Relationships in Picea jezoensis (Pinaceae) Populations of the Russian Far East

Genetic variation and differentiation of 12 populations of Picea jezoensis from the Russian Far East were studied using 20 allozyme loci. The mean number of alleles per locus was 2.63, the percent of polymorphic loci was 88.1%, the observed heterozygosity was 0.181, and the mean value of expected heterozygosity amounted to 0.189. The values of expected heterozygosity of the northern and central mainland populations were higher than in the southern part of the natural range. A significant bias of Hardy–Weinberg heterozygosity to equilibrium heterozygosity (H_eq) suggests that most of the mainland populations have recently experienced a severe expansion in population size while populations from Kamchatka Peninsula have undergone a reduction in population size. Unbiased Nei’s genetic distance values were low within and between the mainland and Sakhalin Island populations (D_N=0.008). The largest values (D_N=0.063) were found between the mainland/Sakhalin and Kamchatka Peninsula populations. Based on genetic distance, P. jezoensis and P. kamtschatkensis could be considered as distinct taxa, but P. ajanensis, P. microsperma, and P. komarovii do not warrant taxonomic recognition.     

Allozyme Diversity and Population Genetic Structure of Pinus densata Master in Northwestern Yunnan, China

We investigated the levels and patterns of genetic diversity of Pinus densata Master in Yunnan. Horizontal starch-gel electrophoresis was performed on macrogametophytes collected from nine populations in northwestern Yunnan, China. Compared with other gymnosperm species, P. densata has higher mean values for all measures of genetic diversity. Allozyme polymorphism (0.99 criterion) was 97.0% and 71.4% at the species and population levels, respectively. The average number of alleles per locus was 3.1 and 2.0 at the species and population levels. Mean expected heterozygosity was substantially higher in P. densata than average values investigated for other gymnosperms both at the population (H _ep = 0.174±0.031) and at the species (H _es = 0.190) levels. Of the total genetic variation, less than 12% was partitioned among populations (G _ST = 0.112). Our allozyme survey supports the suggestion that the observed higher diversity in P. densata may be attributed partly to its hybrid origin between two genetically distinct species, P. yunnanensis and P. tabulaeformis. In addition, we suggest that introgression would give rise to the increase in genetic diversity occurring in P. densata.     

Polymorphic microsatellite loci for population studies of the razor clam, <Emphasis Type="Italic">Sinonovacula constricta</Emphasis>

The razor clam (Sinonovacula constricta) is an important aquacultured bivalve in China. The natural populations of this species are decreasing quickly. To facilitate studies on genetic diversity and population structure of wild populations, microsatellites were isolated from a CA enriched genomic library. Eight microsatellite loci were polymorphic in 30 individuals from Chongming in Shanghai, China. The number of alleles per polymorphic locus varied from 6 to 13 and the values of observed heterozygosity and expected heterozygosity ranged from 0.350 to 1.000 and from 0.602 to 0.902, respectively. These microsatellites are being used in studying population differentiation and genetic diversity for effective conservation and management genetic resources of S. constricta.     

Genetic Variation in Remnant Populations of the Woolly Spider Monkey (Brachyteles arachnoides)

The muriqui or woolly spider monkey (Brachyteles arachnoids) is an endangered primate endemic to the Atlantic Forest of Brazil, <5% of which remains. The known muriqui population consists of <700 individuals separated into approximately 15 geographically isolated forest fragments. I present data on the distribution of genetic variation within and between two such remnant populations (FE and FBR) and summarize the implications of these results for long-range management of species genetic diversity. Eleven of 32 allozyme loci were polymorphic, representing an overall level of polymorphism of 34.4% and a mean heterozygosity per locus of 11%. Both values are among the highest reported for New World monkeys. Genetic differentiation between the two localities is highly significant (F_ST = 0.413, p < 0.001). Genetic distance between them is an order of magnitude greater than that between other populations of platyrrhine subspecies, but this could be an artifact of the small sample size from FBR. High levels of genetic diversity apparently characteristic of this species persist because (1) fragmentation and size reduction of muriqui populations has occurred very rapidly relative to the muriqui life span—although both polymorphism and heterozygosity were lost between generations in the largest population, the high genetic diversity present in the parent population was still in evidence; and (2) genetic diversity before population fragmentation by human activity was not distributed uniformly throughout the species' historic distribution. Thus, remnant muriqui populations are important genetic reservoirs of alleles that are unique or rare in the species gene pool as a whole. These results emphasize the need for the integration of conservation management efforts throughout the species range.     

Isolation and characterization of microsatellite loci of Lycopodium fordii Bak. (Lycopodiaceae, Pteridophyta)

Lycopodium fordii Bak. (Lycopodiaceae, Pteridophyta) is a gardening plant with a native, fragmentary distribution in Taiwan. In this study, we described the development of eleven microsatellite loci in L. fordii for genetic studies. These new markers were tested in 16 individuals of three populations. The number of alleles ranged from 1 to 5 and the expected heterozygosity from 0.41746 to 0.72222. Four of the nine polymorphic loci were significantly deviated from Hardy-Weinberg expectations due to the heterozygote deficiency. The microsatellite markers have also been proved as informative genetic markers for other 15 Lycopodium species.     

A cline of allozyme variation inAbies mariesii

Genetic variation at 22 allozyme loci was examined for 1,003 trees from 11 isolated natural populations ofAbies mariesii covering all except the southernmost region of its geographic range. Genetic diversity within species (H _es=0.063) was low compared to many other long-lived woody species. Most of the genetic variation is found within populations (G _ST=0.144) despite their isolated distribution. Genetic distance between populations was positively correlated with geographic distance. Genetic diversity within populations was generally low (meanH _ep=0.054), but varied across populations in a clinal fashion such that genetic variation decreased with increasing latitude. These genetic characteristics may reflect the distribution history of this species.     

Effects of transgenic American chestnut leaf litter on growth and survival of wood frog larvae

Biotechnology offers a new approach for the restoration of tree species affected by exotic pathogens; however, nontarget impacts of this novel strategy on other organisms have not been comprehensively assessed. We evaluated the effect of transgenic American chestnut (Castanea dentata) leaf litter on the growth and survival of larval wood frogs (Lithobates sylvaticus), a forest‐dwelling amphibian species widely sympatric with American chestnut, that forage almost entirely on periphyton and litter detritus that accumulate in temporary vernal pools in forests. We reared wood frog larvae on Castanea leaf litter (American chestnut genetically engineered for blight tolerance, nontransgenic American chestnut, Chinese chestnut [Castanea mollissima], and an American–Chinese chestnut hybrid) and litter from two non‐Castanea, nontransgenic “control” tree species, coupled with two levels of supplementary food. We observed no differences in growth or survival of wood frog larvae reared on transgenic versus nontransgenic American chestnut leaves. Without supplementary food, wood frog larvae provided leaves from American chestnut (both types) developed faster and grew larger than those exposed to other leaf litter treatments. Results of this study provide preliminary evidence that (1) American chestnut may have formerly been an important source of food for forest‐dwelling amphibians and (2) transgenic American chestnut litter generated as part of chestnut restoration efforts is unlikely to present direct novel risks to developing amphibian larvae in the forest environment.