黄瓜多样性固定核心样本集的构建与评价

利用149对具有多态性的In Del引物对473份黄瓜初选核心种质自交系进行遗传多样性分析。采用3种方法 12种取样比例对该初级遗传多样性固定群体进行抽样获得候选多样性固定核心样本集(GDFCC),使用等位基因数(Na)、有效等位基因数(Ne)、Shannon's信息指数(I)、基因多样性指数(gene diversity)、多态性信息含量(PIC)、总等位位点数(total number of loci)、等位位点保留百分率(retention rate of loci)评价候选多样性固定核心样本集的多样性和代表性,结果表明,采用逐级聚类+稀有基因优先取样法并按照15%取样比例构建出的多样性固定核心样本集的效果较好。比较发现,该核心样本集的Ne、I、基因多样性指数和PIC值均接近或高于初级遗传多样性固定群体,且对原始群体的等位位点的保留百分率为99.68%。入选多样性固定核心样本集的材料来自15个国家和国内18个省市。该研究为今后黄瓜优异基因资源的挖掘利用提供了代表性强、覆盖度广、遗传稳定的研究群体,将有利于黄瓜种质资源的高效研究利用。     

利用RAPD和ISSR分子标记分析地黄种质遗传多样性

用RAPD与ISSR技术对地黄的8个品种和2个脱毒品系进行了种质遗传多样性分析.分别从80条RAPD引物和44条ISSR引物中筛选出适合地黄种质分析的17条RAPD引物和10条ISSR引物用于RAPD和ISSR分析.17条RAPD引物共扩增出177条带, 多态性位点数为109; 多态性位点比率为61.58%;平均多样性指数(I)为0.3135;每个位点的有效等位基因数(Ne)是1.3641; 10条ISSR引物共扩增出110条带. 多态性位点数为79; 多态性位点比率为71.58%;平均多样性指数(I)为0.3577;每个位点的有效等位基因数(Ne)是1.4037. 基于扩增条带数据库建立了各自的Jaccard遗传相关系数矩阵,构建了相似的分子树状图,将10个供试材料分为2类:一类群含组培85.5、大田85.5、组培9302、大田9302、金状元和金白6个材料;另一类群含北京1号、大红袍、地黄9104和野生地黄4个材料.两种分子标记的分析结果呈极显著正相关(r＝0.649).结果表明,RAPD与ISSR标记适合于地黄种质遗传多样性分析,ISSR标记技术是一种多态性和重复性优于RAPD技术的实用技术.     

基于SSR分子标记构建南方型美洲黑杨初级核心种质

以所收集的363份美洲黑杨无性系为实验材料,利用20对SSR引物扩增数据,设置10%、15%、20%、25%、30%、35%、40%共7个取样梯度构建南方型美洲黑杨初级核心种质,并采用平均观察等位基因数(Na)等遗传参数的t检验对核心种质有效性进行确认,以初级核心种质、保留种质以及原始种质三者之间平均观察等位基因数(Na)等遗传参数的保留比例作为评价指标,结合主坐标轴分析法对初级核心种质做进一步代表性确认。结果显示:(1)20对SSR引物共检测到278个多态位点,遗传多样性参数I、H分别为1.667和0.688 2,表明南方型美洲黑杨种质资源具有丰富的遗传变异。(2)比较平均Na、I、H变化确定15%为最佳取样比例,得到54份核心种质和309份保留种质,构建的初级核心种质与原始种质的平均Na、I、H经t检验不显着,除平均Na外其他平均遗传多样性参数,如Ne、Ho、I、H的保留率分别为111%、105%、104%和104%。(3)PCoA分析显示,所构建的初级核心种质不仅与原始种质分布相似,而且分散均匀覆盖较全面,可以代表原始种质在图中的几何分布。研究认为,所构建的45份初级核心种质保存了原始种质大部分的等位基因和基因型,剔除了原始种质的遗传冗余度和遗传重复度,具有更高的遗传多样性参数保留率,能全面代表原始种质的遗传多样性。     

山药种质资源的ISSR分析及初级核心种质库的构建

采用ISSR分子标记技术对中国不同产地的35份山药种质资源进行遗传多样性分析,并用最小遗传距离逐步抽样法构建核心种质库。结果表明:(1)筛选出12个有效引物共扩增出142个位点,多态性比率为97.18%,平均Shannon’s信息指数(I)为0.4230,平均Nei’s基因多样性指数(h)为0.269 4,平均有效等位基因数(Ne)为1.427 1,平均等位基因数(Na)为1.971 8,说明35份山药种质遗传多样性很丰富。(2)聚类分析表明,种质间的遗传相似系数介于0.54~0.97之间,其中来源地不同的个别种质间的遗传相似系数也很高。(3)经6次逐步抽样,随着抽取种质数目的减少,种质库遗传多样性参数变化不明显,而多态性比率在其中5次逐步抽样中呈现下降趋势;但抽样4在抽样数是抽样前的31%时,多态位点率仍可达到抽样前的97.8%,说明抽样4所构建的山药核心种质库最具代表性。     

中型狼尾草种质资源遗传多样性的ISSR分析

运用ISSR标记对采自云南的25份野生种质和4份驯化新品系中型狼尾草材料进行遗传多样性分析。结果显示:(1)50条ISSR引物中共筛选出10条能扩增出清晰条带且多态性明显的引物,29份材料DNA共获得72个扩增位点,其中多态性位点62个,多态性比率为87.4%,平均每条引物扩增位点为7.2个;平均观察等位基因数(Na)、有效等位基因数(Ne)、Shannon多样性信息指数(I)和Nei’s基因多样性指数(H)分别为1.861 1、1.742 8、0.561 0和0.395 9;种质材料间的遗传相似性系数变幅为0.236~0.903,表现出丰富的遗传多样性。(2)利用UPGMA聚类分析,以遗传相似系数0.51为界,29份材料划分为4大类,但Mantel检测表明29份种质材料的遗传聚类和地理距离之间不存在显著的正相关关系(r=0.437 0,P=0.204 6)。研究结果首次从分子水平揭示了中型狼尾草的遗传多样性和变异水平,为合理地引种、驯化、保护和利用中型狼尾草野生资源提供了重要的参考依据和数据支持。     

中国灌木辣椒种质遗传多样性的SRAP和SSR分析

应用SRAP和SSR分子标记对8份辣椒种质进行了遗传多样性分析,结果表明,15对SRAP引物组合共扩增出321条带,平均每对引物扩增出21.40条,多态性位点比率为72.90%;18对SSR引物共扩增出109条带,平均每对引物扩增出6.06条,多态性位点比率为98.17%。与SRAP比较,SSR检测到的Shannon多样性指数(I)、观测等位基因数(Na)和有效等位基因数(Ne)等遗传多样性参数都较大,说明SSR有更高的多态性检测效率。基于SRAP的聚类与基于SSR的聚类之间存在极显著正相关,且都能将中国灌木辣椒种质与美洲灌木辣椒种质及一年生辣椒种质有效区分。     

福建省枳椇种质资源遗传多样性RAPD分析

建立和优化枳椇Hovenia acerba的RAPD反应体系,并对福建省12个种源地35份枳椇材料进行遗传多样性及亲缘关系研究。利用筛选出的10个RAPD引物共检测基因组DNA中193个位点,其中多态性位点161个(83.42%)。供试材料观测等位基因数(Na)1.8342、有效等位基因数(Ne)1.4155,Nei’s基因多样性(H)0.2493,Shannon多样性指数(I)0.3822。以遗传相似系数0.754为阈值,可将35份枳椇分成五大组。RAPD标记可有效揭示福建省枳椇种质资源的遗传多样性,所测枳椇种质资源遗传多样性丰富,极具开发利用价值。     

粤东橄榄资源核心种质取样方案的研究

为研究基于分子标记数据构建橄榄(Canarium album L.)核心种质的取样方案,以粤东地区64份橄榄种质材料的ISSR分析结果为基础,分别利用SM和Nei&Li遗传距离,采用UPGMA聚类法进行多次聚类随机取样,比较了不同分组情况下P、S、L和G等取样策略对核心种质构建的影响。结果表明,通过比较不同样品群的多态性位点数、多态性位点百分率、观测等位基因数、有效等位基因数、Nei’s遗传多样性指数和Shannon’s信息指数等参数,最终选择根据Nei&Li法计算遗传距离,G策略取样得到的16个样品作为核心种质。该核心种质保留了初始种质25%的样品,多态性位点和多态性位点百分率保留率分别为92.93%和98.31%,观测等位基因数、有效等位基因数、Nei’s遗传多样性指数、Shannon’s信息指数的保留率分别为9.26%、102.56%、107.39%和106.29%。因此,按该方案进行取样的核心种质可以较好地代表原有种质库的遗传多样性。     

福建省枳椇种质资源遗传多样性的ISSR分析

通过建立和优化枳椇Hovenia acerba的ISSR反应体系,并利用ISSR分子标记方法,对福建省12个种源地35份枳椇材料进行遗传多样性及亲缘关系研究。枳椇遗传多样性分析的ISSR最佳反应体系为反应总体积25 μL,10×Ex Taq Buffer(Mg2+ plus) 2.5 μL,dNTP Mixture 375 μmol·L-1,引物0.4 μmol·L-1,DNA模板75 ng,Ex Taq酶1 U,ddH2O 14.55 μL。程序：预变性94 ℃ 5 min;变性 94 ℃ 30 s,退火50 ℃ 1 min,延伸72 ℃ 90 s,共 35 个循环;最后延伸72 ℃ 7 min。利用筛选出的10个ISSR引物共检测了基因组DNA中175个位点,其中多态性位点155个,占总位点数的88.57%。供试材料观测等位基因数(Na) 1.8857、有效等位基因数(Ne) 1.4300,Nei’s基因多样性指数(H) 0.2572,Shannon多样性指数(I) 0.3959。UPGMA聚类分析表明,以遗传相似系数0.74为阈值,可将35份枳椇分成四大组。ISSR标记有效地揭示福建省枳椇种质资源丰富的遗传多样性,具较大开发利用价值。     

山茱萸种质资源的ISSR遗传多样性分析与初级核心种质库的构建

采用ISSR 标记技术对不同来源地的48份山茱萸种质资源的遗传多样性进行了分析,并依据最小遗传距离逐步抽样法构建了该初级核心种质资源库.结果显示:筛选出的13个多态性引物共扩增出190个位点,多态性位点比率达93.16%,平均Shannon信息指数(I)为0.402 5,平均Nei's基因多样性指数(H)为0.259 4,平均有效等位基因数(NE)为1.425 0.聚类分析表明,种质间的遗传相似系数介于0.65～0.90之间,除个别种质外,48个种质聚类结果与地区来源有较高的一致性.随着抽取种质数目的减少,多态性比率明显降低,但种质库遗传多样性参数变化较小;抽样3构建的初级核心种质库最具代表性,抽样数是抽样前的30%左右,多态位点比率是抽样前的96.0%.     

黄芩种质资源ISSR遗传多样性的分析及评价

采用ISSR分子标记技术对6个野生或栽培居群共147份黄芩种质进行遗传多样性分析和评价。分析结果表明,51个ISSR引物中筛选出18条扩增条带清晰、重复性好和多态性高的引物,共扩增出485条清晰的条带,其中466条具有多态性,平均多态性位点比率为96.08%,平均Nei’s基因多样性指数和Shannon’s信息指数分别为0.244 4和0.388 9,等位基因数（Na）和有效等位基因数（Ne）分别为1.993 8和1.383 9,遗传分化指数G_st=0.122 3,遗传一致度（I）和遗传距离（D）分别为0.951 5和0.050 1,说明收集的黄芩种质资源在总体上具有较高的遗传多样性,不同居群间存在一定的遗传分化和基因交流,遗传变异主要存在于居群内。分子聚类结果表明,同一地区的种质并没有按照收集来源完全聚类,可能与种质不同起源或民间栽培引种有关。在DNA分子水平揭示黄芩种质资源的遗传多样性水平,将为进一步黄芩种质资源评价、保存和新品种选育等利用提供依据。     

不同地理种源杉木的分子多态性分析

利用ISSR分子标记技术和筛选出的22条ISSR引物,对24个不同地理种源杉木的DNA进行扩增,共扩增出188条谱带,其中多态条带173条,占总数的92.0%.经计算,平均位点的有效等位基因是13408,Nei's基因多样性指数是0.2154,Shannon多态性信息指数为0.3458,表明不同杉木种源间具有较高的遗传多样性.通过UPGMA法聚类分析,可把24个种源杉木分为五个类群:中带东区生态型、中带东南区生态型、中带中区生态型、南带生态型和北带生态型,表明杉木地理种源遗传距离聚类呈现出一定的地域性分布规律.     

Assessment of Genetic Diversity in Three Subsets Constituted from the ICRISAT Sorghum Collection Using Random vs Non-Random Sampling Procedures B. Using molecular markers

The large size of the sorghum [Sorghum bi-color (L.) Moench] landrace collection maintained by ICRISAT lead to the establishment of a core collection. Thus, three subsets of around 200 accessions were established from: (1) a random sampling after stratification of the entire landrace collection (L), (2) a selective sampling based on quantitative characters (PCS), and (3) a selection based on the geographical origin of landraces and the traits under farmers’ selection (T). An assessment was done of the genetic diversity retained by each sampling strategy using the polymorphisms at 15 microsatellite loci. The landraces of each subset were genotyped with three multiplex polymerase chain reactions (PCRs) of five fluorescent primer-pairs each with semi-automated allele sizing. The average allelic richness for each subset was equivalent (16.1, 16.3 and 15.4 alleles per locus for the subsets PCS, L, and T, respectively). The average genetic diversity was also comparable for the three subsets (0.81, 0.77 and 0.80 for the subsets PCS, L, and T, respectively). Allelic frequency distribution for each subset was compared with a chi-square test but few significant differences were observed. A high percentage of rare alleles (71 to 76% of 206 total rare alleles) was maintained in the three subsets. The global molecular diversity retained in each subset was not affected by a sampling procedure based upon phenotypic characters.     

长江春大豆核心种质构建及分析

利用长江春大豆初选核心种质SSR(simple sequence repeat）标记和农艺性状表型等基础数据,对用不同个体取样方法以及不同数据类型建立的核心种质进行评价,目的是确定中国大豆（Glycine max）核心种质的最佳取样策略提供依据,结果表明,根据SSR分子数据聚类,采用类内随机取样,类内以遗传相似性系数取样以及仅依据遗传相似性系数取样都可用于大豆核心种质构建,但是综合不同评价参数发现,以类内随机取样最佳,类内按遗传相似性系数取样次之,单独以遗传相似性系数取样较差。分析不同SSR等位变异保留比例的遗传多样性指数发现,当保留90%和80%的SSR等位变异时,核心种质具有更高的遗传多样性,由于与SSR分子数据种质遗传关系评价的不一致性,农艺性状等基础数据虽然可用来构建核心种质,但其SSR分子水平代表性相对较低,本研究结果还表明,用不同方法或同一方法不同重复次数取样建立的核心种质具有异质性,且这种异质性随核心种质取样比例的降低而增大,因此,虽然可依据不同数据类型确定相应的方法建立核心种质,但综合表型和分子数据建立的核心种质更具有代表性。     

Establishment of a Core Collection of Traditional Cuban <Emphasis Type="Italic">Theobroma cacao</Emphasis> Plants for Conservation and Utilization Purposes

Presently, Theobroma cacao L. (cacao) in Cuba is mainly cultivated in the eastern region where plantations comprise a mixture of clonal varieties, hybrids, progeny of Trinidad Selected Hybrids, and traditional—also known as ancient—cacao. The ancient genetic resources, probably the plants most closely related to the original introductions, are endangered by their progressive replacement by modern clones. To promote the conservation and utilization of these genetic resources, a representative sample of 537 traditional Cuban cacao plants was used to develop a core collection. Core collections based on 15 simple sequence repeat (SSR) markers were generated using five different sampling algorithms: random sampling, simulated annealing, stepwise clustering with random sampling, the M strategy, and maximum genetic diversity. The five core collections were designed to capture 95 % of the SSR alleles in the complete collection. The genetic, morphological, and geographical diversity of each core collection was compared with that of the entire collection. The entire collection contained 139 alleles, including 104 rare ones, with the 95 % allelic coverage threshold achieved with 133 alleles. The core collection generated by the maximum genetic diversity algorithm had the lowest number of accessions (185), the highest mean genetic distance (0.486), the lowest morphological character redundancy, and the highest diversity as assessed by the mean Shannon-Weaver diversity index (0.757). This core collection can thus serve as the basis of future improvement programs based on local genetic resources.     

Assessment of genetic diversity in Salvadora persica L. based on inter simple sequence repeat (ISSR) genetic marker

Studies on the genetic variation in marginal populations and differentiation between them are essential for assessment of best gene conservation strategies and sampling schemes. In this study, ISSR markers were used to establish the level of genetic relationships and polymorphism 50 genotypes of Salvadora persica collected from 6 different regions of Hormozgan province. The ISSR analysis with 9 anchored primers also generated 105 scorable loci, of which 85 were polymorphic (80.95%). Parameters of genetic diversity and its partitioning were calculated. The genetic analysis demonstrated that S. persica maintain relatively high genetic diversity (PIC was 0.63, Na was 1.27 and Ho and He were 0.15 and 0.17 respectively). The coefficient of genetic differentiation among populations based on FST equaled 0.20. Genetic identities between population's pairs were high (mean I?=?0.88). These values are high as compared with other widespread congener species. Cluster analysis based on the Unweighted Pair Group Method with Arithmetic Averages (UPGMA) revealed 3 main clusters for the ISSR data. The levels of genetic diversity maintained within populations of S. persica indicate that an appropriate sampling design for ex situ safeguarding should capture the majority of genetic diversity found within these taxa to help ensure the long term viability of this species. Furthermore, it could be inferred that ISSR markers are suitable tools for the evaluation of genetic diversity and relationships within the Salvadora persica.     

Population structure and genetic diversity in bottle gourd [Lagenaria siceraria (Mol.) Standl.] germplasm from India assessed by ISSR markers

Genetic diversity analysis was undertaken in 42 geographically distant genotypes accessions of bottle gourd (Lagenaria siceraria) from India northeastern (14) and northern region (28) using inter-simple sequence repeat (ISSR) markers. A total of 209 amplified bands were obtained from 20 ISSR primers used in this study, of which 186 were polymorphic with 89.00 % band polymorphism. Various parameters namely, observed number of alleles, effective number of alleles, Nei’s gene diversity/heterozygosity, resolving power, Shannon’s information index and gene flow were estimated under experiment. Jaccard’s similarity coefficient matrix was generated for pairwise comparisons between individual ISSR profiles and UPGMA cluster analysis based on this matrix showed clustering into six groups. Jaccard’s coefficient of similarity values ranged from 0.409 to 0.847, with a mean of 0.628 revealing a moderate level of genetic diversity. The Bayesian model-based approach to infer hidden genetic population structures using the multilocus ISSR markers revealed two populations among the 42 genotypes. This is the first report on the assessment of genetic variation using ISSR markers in this medicinal vegetable plant, and this study of diversity analysis will be helpful in analyzing future hybrid breeding strategy and devising effective germplasm exploration and conservation strategy.     

Optimal sampling strategy and core collection size of Andean tetraploid potato based on isozyme data - a simulation study

Selection of an appropriate sampling strategy is an important prerequisite to establish core collections of appropriate size in order to adequately represent the genetic spectrum and maximally capture the genetic diversity in available crop collections. We developed a simulation approach to identify an optimal sampling strategy and core-collection size, using isozyme data from a CIP germplasm collection on an Andean tetraploid potato. Five sampling strategies, constant (C), proportional (P), logarithmic (L), square-root (S) and random (R), were tested on isozyme data from 9,396 Andean tetraploid potato accessions characterized for nine isozyme loci having a total of 38 alleles. The 9,396 accessions, though comprising 2,379 morphologically distinct accessions, were found to represent 1,910 genetically distinct groups of accessions for the nine isozyme loci using a sort-and-duplicate-search algorithm. From each group, one accession was randomly selected to form a genetically refined entire collection (GREC) of size 1,910. The GREC was used to test the five sampling strategies. To assess the behavior of the results in repeated sampling, k = 1,500 and 5,000 independent random samples (without replacement) of admissible sizes n = 50(50)1,000 for each strategy were drawn from GREC. Allele frequencies (AF) for the 38 alleles and locus heterozygosity (LH) for the nine loci were estimated for each sample. The goodness of fit of samples AF and LH with those from GREC was tested using the L² test. A core collection of size n = 600, selected using either the P or the R sampling strategy, was found adequately to represent the GREC for both AF and LH. As similar results were obtained at k = 1,500 and 5,000, it seems adequate to draw 1,500 independent random samples of different sizes to test the behavior of different sampling strategies in order to identify an appropriate sampling approach, as well as to determine an optimal core collection size.