SSR分子标记在山茶属观赏资源遗传多样性研究中的应用

采用微卫星(SSR)分子标记的方法,利用20对SSR引物对33份山茶属资源(含种和品种)的DNA样品进行了PCR扩增,从中筛选出10对扩增效果较好的SSR引物,并对33份资源进行了遗传多样性分析。结果显示:10对引物在33个植物材料中共检测出123个等位基因,每个SSR位点的等位基因数为3~22个,SSR2引物检测到的等位基因数量最多,达22个,平均每对引物含有12.3个等位基因,平均多态性信息量为0.74,变化范围为0.06~0.96。利用遗传距离矩阵按UPGMA方法进行聚类,结果表明33份植物材料可分为9个分支,很好的区分了品种间的亲缘关系,可为杂交育种的组合选择提供理论基础。不同花型的山茶属植物在10个微卫星位点上共发现42个特异等位基因,可能与不同的花型性状有关。     

陕西省有色稻资源的SSR多态性分析

从水稻12条染色体上筛选出22对有效引物,对陕西省17个有色稻品种的遗传多样性进行分析,共检测到128个等位基因,平均每个标记检测到5.8个等位基因,每个SSR位点的遗传多态性信息含量在0.49-0.89之间,平均值为0.73。聚类分析表明,陕西省17个有色稻品种的遗传相似系数集中在0.24-0.88之间。     

香蕉33个品种的RAPD研究

利用RAPD技术对香蕉（Musa nana Lour.)33个品种的遗传变异进行了研究,从249个随机引物中筛选出18个有效引物,用它们共扩增出192条DNA带,其中183条为多态性带,占95．31％,平均每个引物扩增的DNA带数为10．67条,利用18个有效引物扩增的192条DNA带对香蕉33个品种间的亲缘关系进行UPGMA聚类分析,计算出33个品种间的平均遗传距离为0．3412。在此基础上建立了香蕉33个品种的DNA分子系统树状图。该系统将香蕉33个品种划归A,B,C和D4个群,其中A群20个品种,B群5个品种,C群2个品种,D群6个品种;A群又可以分为3个亚群。对香蕉遗传多样性分子基础进行了探讨。     

鳜原种群体和养殖群体遗传多样性的微卫星分析

本研究利用20对微卫星引物对鳜(Siniperca chuatsi)原种群体和养殖群体进行遗传多样性分析。结果表明,在鳜原种群体中检测到多态性位点14个,养殖群体11个。在两个群体中共检测到等位基因数96个,其中原种群体检测到等位基因数53个,每个位点的等位基因数在1～7之间,平均有效等位基因数为2.7390;养殖群体检测到等位基因数43个,每个位点的等位基因数在1～6之间,平均有效等位基因数为2.1284。原种群体的平均观察杂合度0.5708,Nei氏期望杂合度0.5295,平均多态信息含量PIC0.5353;养殖群体的平均观察杂合度0.3839,Nei氏期望杂合度0.4011,平均多态信息含量PIC0.5043。因此,与养殖群体相比,鳜原种群体仍有丰富的遗传多样性。本研究可为鳜种质资源的保护、监测和遗传育种提供分子水平上的数据。     

利用SSR分析山西省玉米地方品种的遗传多样性

采用混合取样方法和SSR分子标记技术,利用48对引物对山西省38个玉米地方品种的遗传多样性进行了分析.共检测出368个等位基因,每个SSR位点的等位基因数为2～14个,平均为7.48个;多态性信息量(PIC)变化范围在0.24～0.89之间,平均为0.66.总共检测出185个稀有等位基因,21个特有等位基因.SSR标记聚类分析把38个品种大体分成了4个群.研究表明,山西地方品种遗传多样性非常丰富,很多品种具有频率很高的独特基因,它们可能具有一定的特异性.因而,山西玉米地方品种对于拓宽玉米种质的遗传基础可能会起很大的作用.     

东平湖麦穗鱼群体遗传结构的微卫星标记分析

利用微卫星标记技术,采用26对鲤微卫星引物对山东东平湖麦穗鱼进行全基因组扫描.结果表明,有13对引物能获得稳定的特异性条带(占总数的50%),其中有6个微卫星位点具有多态性(占总数的23.1%).6个多态位点共检测到22个等位基因,每个位点的等位基因数从2个到7个不等,大小在80～406bp之间;平均多态信息含量(PIC)为0.5115,平均观测杂合度(H0)为0.6812,平均期望杂合度(HE)为0.5775.研究结果表明,东平湖麦穗鱼群体遗传多样性较丰富,种群结构合理,种质资源处于安全状态.     

不同国家水稻品种的遗传多样性分析

探讨世界不同国家水稻品种的遗传多样性,旨在为各国品种资源的有效利用提供理论依据。本研究利用63对引物对36份来源于不同国家的水稻品种进行遗传多样性分析。共检测到269个等位基因,每个位点的等位基因数(Na)平均为4.54个,有效等位基因数(Ne)平均为3.22,基因多样性指数(H)平均为0.64,Shannon’s信息指数(I)平均为1.21,引物RM206、RM257、RM410、RM235、RM266的等位基因数较多在7条以上。所处纬度相近的国家或地区的水稻品种之间的遗传距离较近,被聚为同一类群,而所处纬度较远的国家或地区的水稻品种被分到了不同类群。结果表明,水稻品种之间的遗传差异与纬度和地理距离有很大的关系。     

平欧杂种榛主栽品种（系）遗传关系的ISSR分析

采用ISSR分子标记的方法,对达维、辽榛3号等17个平欧杂种榛主栽品种(系)进行了遗传关系研究。试验从60条ISSR引物中筛选出7条进行电泳分析,使用NTSYS pc 2.11F和Popgene 1.32软件进行数据统计分析并作图。结果显示:7条ISSR引物共获得58条谱带,平均每条引物8.3条谱带,引物平均多态性比率84.6%;7条ISSR引物可将所有样品完全分开,当遗传相似性阈值为0.687时可将样品分成3个类群:第Ⅰ类群14个品种(系)可分成4个亚类,第Ⅱ类群1个品种,第Ⅲ类群2个品系。样品间的遗传相似性系数为0.448~0.879(平均0.678)。群体的有效等位基因数、基因多样度、Shannon信息指数分别为1.6751、0.3701和0.5308。上述结果表明,平欧杂种榛是一个具有高度遗传多样性的群体,主栽品种(系)间存在复杂的亲缘关系。该研究结果对于平欧杂种榛的遗传关系研究具有重要意义,也可为其他榛属植物的相关研究提供参考。     

利用RAPD和ISSR分子标记分析怀地黄种质遗传多样性

用RAPD与ISSR技术对怀地黄的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引物中筛选出适合怀地黄种质分析的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标记适合于怀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。     

30个粳稻品种SSR标记遗传多样性分析

选用分布于水稻12条染色体上的64对SSR引物,对江苏省育成以及日本引进的粳稻品种共30份材料进行遗传多样性分析。结果表明,有50对SSR引物在30个品种间表现为多态性。共检测到140个等位基因,每对引物的等位基因数变幅为2～5个,平均为2．8个。有效等位基因为94．336个,平均为1．887。每个SSR位点的多态性信息量（PIC）变化范围为0．064～0．752,平均为0．410。30个品种间的遗传相似系数变幅为0．386～0．956之间,平均值为0．719,且81．4％的供试品种其遗传相似系数在0．600～0．800之间,亲缘关系较近;以遗传相似系数为原始数据,按UPGMA方法将30个品种划分为3大类群,结合系谱分析结果表明,江苏省育成的水稻品种遗传多样性不够丰富,多数品种间的亲缘关系较近,欲进一步提高江苏省水稻产量还需拓宽亲本选择范围,扩大遗传背景。     

Genetic diversity of Carica papaya as revealed by AFLP markers.

Genetic relationships among Carica papaya cultivars, breeding lines, unimproved germplasm, and related species were established using amplified fragment length polymorphism (AFLP) markers. Seventy-one papaya accessions and related species were analyzed with nine EcoRI-MseI primer combinations. A total of 186 informative AFLP markers was generated and analyzed. Cluster analysis suggested limited genetic variation in papaya, with an average genetic similarity among 63 papaya accessions of 0.880. Genetic diversity among cultivars derived from the same or similar gene pools was smaller, such as Hawaiian Solo hermaphrodite cultivars and Australian dioecious cultivars with genetic similarity at 0.921 and 0.912, respectively. The results indicated that self-pollinated hermaphrodite cultivars were as variable as open-pollinated dioecious cultivars. Genetic diversity between C. papaya and six other Carica species was also evaluated. Carica papaya shared the least genetic similarity with these species, with an average genetic similarity of 0.432; the average genetic similarity among the six other species was 0.729. The results from AFLP markers provided detailed estimates of the genetic variation within and among papaya cultivars, and supported the notion that C. papaya diverged from the rest of Carica species early in the evolution of this genus.     

Sequence related amplified polymorphism (SRAP) analysis for genetic diversity and micronutrient content among gene pools in mungbean [Vigna radiata (L.) Wilczek]

Vigna radiata (L.) Wilczek, commonly called mungbean is an important pulse crop. Commercial cultivars contain low levels of iron and zinc and it is important to assess genetic variability in the available germplasm for improving micronutrient content in commercial cultivars. The present study was undertaken to study molecular diversity using Sequence-related amplified polymorphism (SRAP) among 21 Vigna radiata genotypes. Twenty nine SRAP primer combinations produced a total of 121 amplified bands which were polymorphic with an average of 4.65 bands per primer. The size of amplified bands ranged from 70 bp to 3,000 bp and 6 out of 29 SRAP primers were most useful in fingerprinting Vigna radiata genotypes under study. The similarity coefficients between different genotypes ranged from 0.45 to 0.96 with an average similarity value of 0.71. At an arbitrary cut-off at 60 % similarity level on a dendrogram, the Vigna radiata accessions were categorized into two major clusters. ML1108 and 2KM115 were found to be genetically similar. SMH99-1A and ML776 showed high iron and zinc content while Satya was poor in iron as well as zinc content. Mapping population involving ML776 and Satya could be used for tagging gene(s) for micronutrient content. The results indicated that SRAP markers were efficient for identification of Vigna radiata genotypes and assessment of the genetic relationships among them.     

Genetic diversity in colonial bentgrass (Agrostis capillaris L.) revealed by EcoRI-MseI and PstI-MseI AFLP markers.

Colonial bentgrass (Agrostis capillaris L.) is a potential source for genetic improvement of resistance to environmental stress and disease for other bentgrass species (Agrostis spp.). To conserve and study the existing genetic resources of colonial bentgrass for use in breeding, genetic diversity was investigated using amplified fragment length polymorphism (AFLP) markers. Included in this study were 22 accessions from US Department of Agriculture germplasm collected from 11 countries, in conjunction with 14 accessions from northern Spain and 3 commercial cultivars. Ten EcoRI-MseI and 6 PstI-MseI AFLP primer combinations produced 181 and 128 informative polymorphic bands, respectively. Cluster analysis of genetic similarity estimates revealed a high level of diversity in colonial bentgrass species with averages of 0.51 (EcoRI-MseI) and 0.63 (PstI-MseI). Greater genetic diversity was detected by the EcoRI-MseI AFLP primer combinations. A low but significant positive correlation (r = 0.44, p = 0.0099) between the 2 Jaccard similarity matrices was obtained by the Mantel test. Commercial cultivars of bentgrass showed a narrow genetic background. The assessment of genetic diversity among colonial bentgrass accessions suggested the potential value of the colonial bentgrass germplasm in turfgrass cultivar improvement.     

Assessment of genetic variation within Indian mustard (Brassica juncea) germplasm using random amplified polymorphic DNA markers

Genetic diversity among 45 Indian mustard (Brassica Juncea L.) genotypes comprising 37 germplasm collections, five advance breeding lines and three improved cultivars was investigated at the DNA level using the random amplified polymorphic DNA (RAPD) technique. Fifteen primers used generated a total of 92 RAPD fragments, of which 81 (88%) were polymorphic. Of these, 13 were unique to accession 'Pak85559'. Each primer produced four to nine amplified products with an average of 6.13 bands per primer. Based on pairwise comparisons of RAPD amplification products, Nei and Li's similarity coefficients were calculated to evaluate the relationships among the accessions. Pairwise similarity indices were higher among the oilseed accessions and cultivars showing narrow ranges of 0.77-0.99. An unweighted pair-group method with arithmetic averages cluster analysis based on these genetic similarities placed most of the collections and oilseed cultivars close to each other, showing a low level of polymorphism between the accessions used. However, the clusters formed by oilseed collections and cultivars were comparatively distinct from that of advanced breeding lines. Genetically, all of the accessions were classified into a few major groups and a number of individual accessions. Advanced breeding lines were relatively divergent from the rest of the accessions and formed independent clusters. Clustering of the accessions did not show any pattern of association between the RAPD markers and the collection sites. A low level of genetic variability of oilseed mustard was attributed to the selection for similar traits and horticultural uses. Perhaps close parentage of these accessions further contributed towards their little diversity. The study demonstrated that RAPD is a simple and fast technique to compare the genetic relationship and pattern of variation among the gene pool of this crop.     

Genetic characterization of Iranian safflower (Carthamus tinctorius) using inter simple sequence repeats (ISSR) markers

Safflower (Carthamus tinctorious L.) is valued as a source of high quality vegetable oil. 20 ISSR primers were used to assess the genetic diversity of 18 accessions of safflower collected from different geographical regions of Iran. The ISSR primers combinations revealed 57.6 % polymorphism, among 338 genetic loci amplified from the accessions. The sum of effective number of alleles and observed number of alleles were 29.76 and 36.77, respectively. To understand genetic relationships among these cultivars, Jacquards’ similarity coefficient and UPGMA clustering algorithm were applied to the ISSR marker data set. ISSR markers grouped accessions into two main clusters and four sub clusters. Also, the principal coordinate analysis (PCoA) supported the cluster analysis results. The results showed these genotypes have high genetic diversity, and can be used for alternative safflower breeding program.     

Molecular characterization and genetic diversity analysis of soybean (Glycine max (L.) Merr.) germplasm accessions in India

Molecular characterization and genetic diversity among 82 soybean accessions was carried out by using 44 simple sequence repeat (SSR) markers. Of the 44 SSR markers used, 40 markers were found polymorphic among 82 soybean accessions. These 40 polymorphic markers produced a total of 119 alleles, of which five were unique alleles and four alleles were rare. The allele number for each SSR locus varied between two to four with an average of 2.97 alleles per marker. Polymorphic information content values of SSRs ranged from 0.101 to 0.742 with an average of 0.477. Jaccard’s similarity coefficient was employed to study the molecular diversity of 82 soybean accessions. The pairwise genetic similarity among 82 soybean accessions varied from 0.28 to 0.90. The dendrogram constructed based on genetic similarities among 82 soybean accessions identified three major clusters. The majority of genotypes including four improved cultivars were grouped in a single subcluster IIIa of cluster III, indicating high genetic resemblance among soybean germplasm collection in India.

Electronic supplementary material

The online version of this article (doi:10.1007/s12298-014-0266-y) contains supplementary material, which is available to authorized users.     

利用RAPD标记分析大麦种质资源的遗传多样性

利用RAPD标记对19份西藏近缘野生大麦材料、33份我国不同省市的地方品种以及8份国外引进大麦品种共60份大麦种质资源的遗传多样性进行检测.结果表明材料间遗传差异明显.32个RAPD引物中,有25个引物(占78.13%)可扩增出清晰且具多态性的条带,另外7个引物能扩增出1～3条清晰但无多态性的条带.每个引物可扩增出1～8条多态性带,平均为3.72条.32个引物共产生119条DNA片段,其中87条具有多态性,多态性比率(PPB)为73.11%,平均多态信息量(PIC)为0.434;每个位点平均有效等位基因数(Ne)为2.304;材料间遗传相似系数GS变化范围为0.757～0.981,平均值为0.871.19份来源于西藏的近缘野生大麦材料间GS值变幅为0.818～0.969,平均为0.892;33份我国栽培大麦地方品种间的GS值变化范围为0.783～0.981,平均为0.879;8份分别来自8个国家的栽培大麦品种间的GS值变幅为0.820～0.956,平均为0.882.根据RAPD标记分析的结果,对60份大麦种质资源进行聚类分析,在平均GS值0.871水平上60份大麦材料可聚为5类,聚类结果能在一定程度上反应材料的地理分布关系,但某些相同地理来源的材料也较分散地分布在整个聚类树中.本研究从分子水平上进一步证明了我国栽培大麦丰富的遗传多样性,是世界栽培大麦的遗传多样性中心之一.     

AFLP analysis of genetic diversity within and among Coffea arabica cultivars

Genetic diversity of Coffea arabica cultivars was estimated using amplified fragment length polymorphism (AFLP) markers. Sixty one Coffea accessions composed of six arabica cultivars, including Typica, Bourbon, Catimor, Catuai, Caturra and Mokka Hybrid, plus two diploid Coffea species, were analyzed with six EcoRI- MseI primer combinations. A total of 274 informative AFLP markers were generated and scored as binary data. These data were analyzed using cluster methods in the software package NTSYSpc. The differences among cultivars at the DNA level were small, with an average genetic similarity of 0.933. Most accessions within a cultivar formed a cluster, although deviant samples occurred in five of the six cultivars examined due to residual heterozygosity from ancestral materials. Among the six cultivars fingerprinted, the highest level of genetic diversity was found within the cultivar Catimor, with an average genetic similarity of 0.880. The lowest level was found within Caturra accessions, with an average genetic similarity of 0.993. Diversity between C. arabica and two other Coffea species, Coffea canephora and Coffea liberica, was also estimated with average genetic similarities of 0.540 and 0.413, respectively, suggesting that C. canephora is more closely related to C. arabica than is C. liberica. The genetic variation among arabica cultivars was similar to the variation within cultivars, and no cultivar-specific DNA marker was detected. Although arabica cultivars appear to have a narrow genetic base, our results show that sufficient polymorphism can be found among some arabica cultivars with a genetic similarity as low as 0.767 for genetic/QTL mapping and breeding. The assessment of genetic diversity among arabica cultivars provided the necessary information to estimate the potential for using marker-assisted breeding for coffee improvement.     

中国食用向日葵种质资源遗传变异的RAPD及AFLP分析

本研究采用RAPD和AFLP方法对23个中国不同地区的食用向日葵（Helianthus annuus L.)骨干品种进行了遗传变异分析,同时对两种标记系统进行了比较。26个RAPD引物产生了总计192条DNA条带,大小分布 于0.26kb-1.98kb之间,其中165条（86．12％）具有多态性,每条引物产生DNA条带的平均数为7．38。8对AFLP引物组合共产生了576条带,分布于100bp-500bp之间,其中的341条具有多态性,多态百分率为76．00％,每对引物组合产生DNA条带的平均数为72。RAPD方法检测的每位点有效等位基因数（1．76）大于AFLP（1．65）,AFLP标记位点的平均多态性信息量（PIC）（0．38）低于RAPD标记位点PIC（0．41）,但AFLP标记具有很高的多态性检测效率（Ai=38．52）。用RAPD标记分析23个食用向日葵材料的亲缘关系,Nei氏相似性系数分布在47．84％－82．06％,平均相似性系数为0．6495,而采用AFLP的Nei氏相似性系数分布在54．15％－83．52％,平均相似性系数为0．6884。RAPD数据的标准差为0．13,而AFLP数据的标准差为0．08。因此,采用RAPD和AFLP方法分析食用向日葵遗传变异,RAPD标记具有较低相似性系数和较高方差而AFLP则相反。源于两种不同标记的遗传相似矩阵的相关系数为0．51,说明采用RAPD和AFLP系统分析食用向日葵遗传变异得到的结果有一定的相关性,无论采用RAPD还是AFLP标记进行聚类分析,都将23个不同基因型的食用向日葵材料分成了三个类群。     

Genetic Diversity of Some Pear Cultivars and Genotypes Using Simple Sequence Repeat (SSR) Markers

The genetic diversity and relationships among 47 pear cultivars and genotypes (Pyrus spp.), including 4 Japanese pears (Pyrus pyrifolia), 40 European pears (Pyrus communis), 1 Chinese pear (Pyrus bretschneideri) as well as 2 wild relatives (Pyrus salicifolia and Pyrus mazandaranica) were studied using 28 microsatellite primer pairs. A total of 174 alleles were produced at the 28 SSR loci with their sizes ranging from 81 to 290?bp. The number of observed alleles for each locus ranged from 3 (TsuENH014 and TsuENH046) to 12 (NB103a), with an average of 6.21 alleles per locus. In some SSR loci, more than two alleles were amplified in some cultivars and genotypes, suggesting that duplication has occurred in those accessions. This information suggests that at least two genomic regions exist for these loci in the pear genome. The observed heterozygosity (H _o) values of amplified loci ranged from 0.17 (TsuENH006) to 0.97 (NB103a). Shannon's information index (I) value was observed to be highest (2.14) in the NB103a locus, while the TsuENH006 locus had the lowest value with an average of 1.37 among SSR loci. The Dice genetic similarity coefficient ranged from 0.29 (??Nijisseiki?? and P. mazandaranica) to 0.91 (??Chojuro?? and ??Nijisseiki??) among samples. UPGMA cluster analysis showed two major groups corresponding to the Japanese and European pears.