Molecular marker diversity and bacterial wilt resistance in wild <Emphasis Type="Italic">Solanum commersonii</Emphasis> accessions from Uruguay

Solanum commersonii is a wild tuber-bearing species native to Uruguay with high potential for use in potato breeding programs. Little is known about the genetic diversity within this wild species and the relationship with the resistance to the bacterial pathogen Ralstonia solanacearum. We studied 30 S. commersonii clonal accessions, 20 of which were collected from geographically different areas across the country, while the other ten were grown from seeds from a single plant. Resistance against R. solanacearum was tested and different levels of resistance were found, ranging from delayed wilting to asymptomatic reactions. The genetic variation and the relationships among individuals in this germplasm collection were studied by different molecular markers: Random Amplified Polymorphic DNA (RAPD), Amplified Fragment Length Polymorphism (AFLP) and Microsatellites or Simple Sequence Repeats (SSR). AFLP markers generated the largest number of total and polymorphic fragments per assay unit while SSR revealed the highest frequency of polymorphic bands (100%), followed by AFLP (96.2%) and RAPD (89.4%). In contrast, when comparing the number of different genetic profiles generated, the SSR markers exhibited the lowest discriminatory power. The clustering pattern obtained with the three marker systems showed a similar distribution of the S. commersonii germplasm revealing a high correlation between the three methods employed. All three dendrograms grouped most of the accessions into two main clusters, containing the same accessions regardless of the marker type. Bacterial wilt resistant accessions were present in both clusters. Accessions originated from different seeds of the same plant were grouped within one of the major clusters, and differed in the response to R. solanacearum revealing segregation of resistance. Furthermore, the distribution in two main clusters showed high correspondence with the geographical origin of the accessions, from the north and south of the country, and with the subspecies malmeanum and commersonii morphologically identified.     

利用SSR标记分析橡胶草种质资源的遗传多样性

为了解橡胶草种质的遗传背景和遗传多样性,为今后橡胶草育种提供理论依据。利用23对SSR引物对96份橡胶草材料进行遗传多样性分析。结果显示,23对SSR引物通过扩增得到71个等位变异,等位变异范围2-6个,平均等位基因数为3.09个。通过聚类分析,俄罗斯材料和美国材料与新疆7个居群材料被分为2大类群,类群I包含所有俄罗斯和美国材料以及5份新疆野生材料,类群II包含其余新疆7个居群的材料;俄罗斯和美国材料同属于亚群A,平均遗传相似度为0.88,说明它们存在紧密的亲缘关系;新疆7个居群的材料被分为5个亚群,显示丰富的遗传多样性,而且相互之间存在复杂的遗传关系。本研究结果证明了SSR标记能够有效地用于橡胶草的遗传多样性研究,为以后的橡胶草种质收集和遗传育种提供重要依据。     

Microsatellite (SSR) variation in a collection of Malus (apple) species and hybrids

A collection of 142 accessions of 23 Malus species, derived hybrids and cultivar accessions from the USDA-ARS Plant Genetic Resources Unit's core collection, which represents an extensive range of Malus species, was screened with a set of previously described SSR (simple sequence repeat) markers. The markers were used to determine genetic identities, estimate genetic diversity, identify genetic relationships among the accessions, and determine the utility of SSR primers developed from Malus ×domestica for making genetic assessments across the whole Malus genus. All eight primer pairs amplified multiple fragments when used in polymerase chain reactions with DNA from these accessions. High levels of variation were detected with a mean of 26.4 alleles per locus and a mean direct count heterozygosity across all eight loci equal to 0.623. The eight primer pairs used in this study unambiguously differentiated all but five pairs of accessions in this collection of 142 accessions of 23 Malus species, derived hybrids and cultivars. These SSR data were not useful in identifying genetic relationships among this diverse collection of accessions, with the majority of the accessions not clustering in ways concordant with taxonomic information and/or geographic origin. The resulting phenogram resolved only two meaningful clusters, for the taxonomically isolated Section Chloromeles and for M. fusca accessions, reflecting genetic relationships arising from geographic origin. The detection of identical accessions in the collection, which were previously considered to be unique, highlights the critical need to further bolster collections of certain Malus species. This revised version was published online in July 2006 with corrections to the Cover Date.     

利用SSR分析四倍体棉种多态性

利用SSR标记对60份四倍体棉种材料进行遗传多样性分析,其中包括42份陆地棉野生种系。结果显示, 从1 050对SSR引物筛选出的95对SSR引物均能在60份材料间扩增出稳定明显的多态性条带,共检测出660个片段,其中多态性片段584个,占88.5%。每个位点的等位基因为2~12个,平均每对引物6.1个。UPGMA聚类分析显示, 42份材料的相似性系数(GS)在0.306~1.000之间,平均成对相似系数为0.493。95对引物的多态信息含量(PIC)的变幅为0.278~0.905,基因多样性(H¢)的变幅为0.451~2.451, 有效等位基因数(Ne)在1.385~10.490之间变动。SSR标记在陆地棉野生种系及四倍体棉种间均可反映丰富的遗传多样性信息,其中陆地棉与陆地棉野生种系中阔叶棉的亲缘关系最近,海岛棉和达尔文棉的亲缘关系非常相近,黄褐棉和毛棉相对较近,陆地棉与其他4个棉种的亲缘关系最远。     

Genetic diversity in Cynara cardunculus revealed by AFLP markers: comparison between cultivars and wild types from Sicily*

Genetic diversity in 49 plants of 16 accessions representing one wild and two cultivated taxa of Cynara cardunculus L. (wild cardoon, cultivated cardoon and globe artichoke) was investigated using amplified fragment length polymorphic (AFLP) analysis. Four pairs of primers identified a total of 264 scorable loci, 196 of which were polymorphic. The number of markers scored per primer combination ranged from 41 to 107, with an average of 66 markers per primer combination. All the primer combinations detected polymorphism. The data were used for cluster analysis and AMOVA. The clusters obtained are in agreement with the current botanical classification and the differences among the taxa were substantial. The eco‐geographical groups within wild cardoon are clearly separated in relative subclusters, and reflect the geography of the collection areas. The six clones of the four cultivated artichoke varieties were clearly separated into four subclusters, corresponding to the current classification which is based on morphological traits. The results obtained may have a direct and important bearing on plant genetic resource conservation and management and on breeding.     

Genetic linkage maps of Citrus sunki Hort. ex. Tan. and Poncirus trifoliata (L.) Raf. and mapping of citrus tristeza virus resistance gene

The RAPD technique was used to identify genetic relationships in 19 accessions, including six species of the genus Chenopodium. A dendrogram was constructed using UPGMA from 399 DNA markers. The molecular data clustered species and accessions into five different groups. Group 1 with three cultivated varieties of C. nuttalliae, Group 2 included eight cultivars and two wild varieties of C. quinoa, Group 3 with C. berlandieri and C. album, Group 4 with two accessions of C. pallidicaule, and Group 5 with 2 accessions of C. ambrosioides. The polymorphic patterns generated by RAPD profiles showed different degrees of genetic relationship among the species studied. A low level of intraspecific variation was found within the accessions of C. quinoa, C. nuttalliae, and C. pallidicaule. The RAPD markers were found to be a useful tool for detecting genetic variation within the genus Chenopodium.     

利用SSR标记分析野生小豆及其近缘野生植物的遗传多样性

利用28对SSR引物对96份野生小豆资源、小豆近缘野生植物及栽培小豆品种进行遗传多样性分析,共检测到255个等位变异,平均每对SSR引物9.10个,多态信息含量的变异范围从0.374到0.865,平均为0.722。野生小豆材料和近缘植物Vigna minima遗传变异丰富。来自不同地域的野生小豆材料具有大量特异等位变异,基于非加权成组配对算术平均法的聚类分析可将不同地理来源的野生小豆单独分群,且与主坐标分析的结果相一致。4份栽培小豆材料与日本野生小豆遗传距离较近,表明目前国内小豆育种中较多使用了含有日本血缘的小豆材料,以及国内野生小豆资源的搜集和利用工作落后于日本。本研究对国内野生小豆资源的搜集和保存具有指导意义,并可以为这些资源的评价、利用和优异基因的发掘提供参考。     

中国普通野生稻与栽培稻种SSR多样性的比较分析

采用48对SSR引物对288份我国普通野生稻和栽培稻的遗传多样性进行比较分析。结果显示, 共检测到505个等位基因, 每个位点的等位基因数变幅为5~20, 平均10.5个; 平均Nei基因多样性指数(He)为0.731, 变幅为0.384(RM409)~0.905(RM206)。普通野生稻遗传多样性高于栽培稻种, 栽培稻等位基因数和平均Nei基因多样性指数分别为普通野生稻的70.2%和88.2%, 其中, 栽培稻地方品种和选育品种等位基因数分别为普通野生稻的65.4%和53.0%, 选育品种等位基因数仅为地方品种的81.1%。AMOVA分析表明, 总变异的10.3%是由于种间SSR遗传差异所引起的, 不同SSR位点种间的分化程度不同, 在0.7%~46.3%之间, 有43个位点种间遗传分化达到显著水平, 其中以RM427分化最为明显, 达46.3%。聚类分析表明, 中国普通野生稻总体偏粳, 极少数广东、海南材料偏籼。     

Random amplified polymorphic DNA (RAPD) variation in Fagopyrum tataricum Gaertn. accessions from China and the Himalayan region

The diversity among 52 landraces and cultivars of tartary buckwheat (Fagopyrum tataricum Gaertn.) and one accession of its wild ancestor, F. tataricum ssp. potanini Batalin, from diverse geographic origins was examined using random amplified polymorphic DNA-polymerase chain reaction (RAPD-PCR) markers. Eighteen primers produced a total of 240 fragments, of which 153 (63.75%) were monomorphic and 87 (36.25%) polymorphic bands. UPGMA-based pairwise Jaccard’s coefficient of similarity was used to deduce the relationships among 53 genetically diverse accessions. The similarity between cultivated tartary buckwheat accessions ranged from 0.61 to 1.00. Four distinct clusters were formed which corresponded well with the geographic distribution of the tartary buckwheat. Nepalese accessions showed maximum diversity followed by Chinese accessions. Tartary buckwheat accessions from the Himalayan region of northwestern India revealed a narrow gene pool. The wild buckwheat accession did not group with any of the three cultivated tartary buckwheat groups, and formed its own single-entry group. Genetic similarity (0.59) of Chinese buckwheat accessions with the wild ancestor reaffirmed that cultivated tartary buckwheat originated in the Yunnan province of northwestern China. Consistent with some earlier reports, our study demonstrated the usefulness of the RAPD technique for the characterization of plant genetic resources and assessment of diversity between species. This revised version was published online in August 2006 with corrections to the Cover Date.     

应用SSR标记对小豆种质资源的遗传多样性分析

利用24对SSR引物对158份栽培小豆及18份野生小豆资源进行遗传多样性分析。结果表明,栽培小豆的遗传变异水平显著低于野生小豆,其中18对引物在栽培小豆中能检测到多态性,平均等位变异数为3.0个,21对引物在野生小豆中能检测到多态性,平均等位变异数为2.6个。栽培小豆种质间平均遗传相似性系数为0.724,野生小豆间为0.605。基于类平均法的聚类分析可以将栽培小豆和野生小豆区分开,这与主坐标分析的结果基本吻合。不同来源的栽培小豆群体间也有一定的遗传分化。SSR分析不仅验证了小豆品种间的遗传背景与其系谱来源相吻合,而且揭示了同名种质天津红小豆之间的遗传差异。本研究为我国小豆种质资源育种及SSR标记在小豆多样性分析、基因标记、品种鉴定等工作提供了信息。     

应用SSR和ISSR标记分析栽培香稻品种的遗传多样性

本研究利用24对SSR引物和36个ISSR引物,分析33份来源于亚洲10个国家的香稻品种的遗传多样性。分别获得93条和181条多态性片段,每个SSR座位可检测3～8个等位基因,平均为4．23个;每个ISSR引物可检测3～8个多态性位点,平均为5．03个。根据SSR和ISSR标记计算的品种间遗传相似系数分别在0．294～0．884之间和0．595～0．867之间。聚类分析表明,利用两种标记所得的聚类结果基本上一致,与品种所处的3种气候类型变化基本相符。进一步证实SSR和ISSR标记是研究水稻种质资源分类有效的工具。     

利用SSR标记分析栽培种花生多态性及亲缘关系

利用11对SSR引物对24个花生栽培品种（包括四大类型）进行PCR扩增分析，其中4对检测到明显的多态性，共检测到33个等位基因变异，每一个位点上检测到的等位变异数为5～13个，平均为8.25个。根据扩增结果可以将24个品种中的21个相互区分。供试品种间的遗传相似系数值在0.2～1.0之间，平均为0.4788。根据UPGMA聚类分析结果,供试     

Genotyping safflower (Carthamus tinctorius) cultivars by DNA fingerprints

Summary Carthamus tinctorius (2n = 2x = 24) (family Asteraceae), commonly known as safflower, is widely cultivated in agricultural production systems of Asia, Europe, Australia and the Americas as a source of high-quality vegetable and industrial oil. India ranks first in the production of safflower oil. Fourteen cultivars, widely cultivated in various agro-climatic regions of India, have been fingerprinted by RAPD, ISSR, and AFLP markers utilizing 36, 21 primers, and 4 primer combinations, respectively. On an individual assay basis, AFLP has proven to be the best marker system as compared with the other two markers applied as assessed by high discriminating power (0.98), assay efficiency index (33.2), marker index (18.2), resolving power (40.62), and genotype index (0.856). Thirty-six RAPD and 21 SSR primers could differentiate a maximum of eight and four cultivars, respectively, whereas, two AFLP primer combinations could fingerprint all the 14 cultivars. To understand genetic relationships among these cultivars, Jaccard's similarity coefficient and UPGMA clustering algorithm were applied to the three marker data sets. Mean genetic similarities ranged from 0.689 (AFLP) to 0.952 (ISSR). Correlation coefficient comparisons between similarity matrices and co-phenetic matrices obtained with the three markers revealed that AFLP displayed no congruence vis-a-vis RAPD and ISSR data. However, strong correlation was observed between RAPD and ISSR marker systems. This paper reports the start of molecular biology programme targeting nuclear genome of safflower, a major world oilseed crop about whose genetics very little is known.     

利用SSR分子标记构建‘太湖糯’、‘苏御糯’及部分优质糯稻的DNA指纹图谱

为了探明国内部分优质糯稻资源的遗传背景,从分子水平揭示其遗传多样性,同时为优质糯稻资源的种质鉴定和创新利用提供理论依据,以太湖稻区地产优质糯稻太湖糯、苏御糯和国内其他地区部分糯稻共14个品种为研究材料,利用SSR分子标记进行了DNA指纹图谱的构建,并进行聚类分析,研究其亲缘关系。结果表明,从35对国标中公布的水稻SSR引物中筛选出17对核心引物,在这14个糯稻品种中共检测到63个多态性片段。利用这17对核心引物进行聚类分析,发现供试14个品种的遗传相似系数在0.619以上,表明14个糯稻品种具有较丰富的遗传变异;并利用其中6个引物,经PCR扩增获得的23条清晰、稳定、重复性好的多态扩增条带,建立了14个糯稻品种的DNA指纹图谱。     

Molecular diversity and genome-wide linkage disequilibrium patterns in a worldwide collection of <Emphasis Type="Italic">Oryza sativa</Emphasis> and its wild relatives

Genetic diversity analysis within a species is vital for understanding evolutionary processes at the population and genomic levels. We report a detailed study of molecular diversity, polymorphism and linkage disequilibrium in three groups of rice (Oryza) germplasm accessions based on 176 SSR markers. The first group included 65 rice (O. sativa L.) accessions introduced from seven countries, including five regions of China. The second group included 58 US rice varieties released in the past 25 years. The third group consisted of 54 accessions of rice wild relatives represented by ten different species. The number of alleles per SSR marker ranged from 4 to 32 with a mean of 16 alleles and the polymorphism information content values ranged from 0.43 to 0.91 with a mean of 0.70. The variation in SSR alleles was a significant contribution to the genetic discrimination of the 177 accessions within the three Oryza groups. Analysis of molecular variance identified deviation from Hardy–Weinberg equilibrium. Principal coordinates analysis clearly separated the accessions into their respective three groups. Neighbor-joining phylogenetic cluster reflects the ordination of each accession. Linkage disequilibrium (D′) averaged 0.75 in wild Oryza spp., and about 0.5 in both US and international O. sativa accessions. Our results showed that LD among adjacent loci in both O. sativa and Oryza spp. accessions is strong enough to be detecting marker-trait association via genome-wide scans.     

Genetic diversity in tef [Eragrostis tef (Zucc) Trotter] and its relatives as revealed by Random Amplified Polymorphic DNAs

Tef is one of the staple cereal crops in Ethiopia. To evaluate genetic diversity of tef and its relatives, 47 accessions of tef, three accessions of E. pilosa, and six accessions of E. curvulawere analyzed using random amplified polymorphic DNA (RAPD) markers. The level of polymorphism among the wild species was extremely high, while low polymorphism was detected among tef accessions. All cultivars and wild species under study could be distinguished with the help of different primers, thereby indicating the potential of RAPD in the genetic fingerprinting of tef. Accessions from E. curvula and E. pilosa can be differentiated by a single selected primer. In spite of low polymorphism within tef, accessions under study could be distinguished by a combination of selected primers. Cluster analysis indicated that tef is a very closely related species to E. pilosa with 45%similarity, supporting the hypothesis that tef originated from E. pilosa based on morphological data. Given that RAPD are relatively quick, simple to use, and are not subjected to environmental influences, they provide a valuable new approach for the genetic fingerprinting and study of genetic diversity in tef. This revised version was published online in July 2006 with corrections to the Cover Date.     

Inter and intra-species Inter Simple Sequence Repeat (ISSR) variations in the genus Cicer

Intra and inter-species ISSR variation and use of ISSR markers in determination of genetic relationship were investigated in an accession collection representing twoperennial and six annual Cicerspecies. Screening of Ciceraccessions with SSR primers revealed highly reproducible amplicon profiles with relatively high multiplex ratios. Many of the primers generated amplicon profiles with which not only the differences among species can readily be identified, but also polymorphisms within species could be detected more efficiently. PCR products at 150 gel positions detected using six SSR primers in Cicer accessions were treated as dominant DNA markers and utilized to compute the distances among accessions and species. Cluster analysis of accessions and species revealed groupings that corroborate our previous studies of relationships based on allozyme and AFLP analysis. Consistent with the AFLP analysis carried out in the same accession collection, ISSR-based groupings indicated that perennial C. incisumis genetically close to the annuals of the second crossability group (C. pinnatifidum,C. bijugum, C. judaicum) while C. reticulatum is the closest wild species to the cultivated chickpea. ISSR-based variation estimates were relatively higher when compared to previous estimates computed from RAPD and AFLP data. Technically, ISSR analysis combines the PCR-based targeting of microsatellite-associated polymorphisms with no prior sequence requirement and stringent PCR conditions. Similarly, when compared to AFLP analysis, it is less technically demanding allowing to survey polymorphic loci in the genome. Thus, ISSR-PCR technology is a reliable, fast, and cost-effective marker system that can be used to study genetic variation and genetic relationships in the genusCicer. This revised version was published online in July 2006 with corrections to the Cover Date.     

36个马铃薯品种的SSR分析

为明确36个马铃薯品种在DNA分子水平上的遗传差异,利用SSR分子标记技术对其多态性进行了分析。试验从90对SSR引物中筛选出适宜于36个马铃薯品种基因组DNA扩增的引物10对,PCR扩增获得301个SSR条带,多态性条带百分率为71.1%。引物STACCA3和SSR111扩增的SSR指纹图谱差异明显,可作为36个马铃薯品种间鉴别的分子依据。36个马铃薯品种间的遗传距离(GD)变幅为0.33～0.85,平均为0.54,其中,大于GD平均值的品种有20个,占供试品种的55.56%。以GD值0.57为基准,将36个马铃薯品种划分成7类,从DNA分子水平揭示出各供试品种间的亲缘关系。     

Comparative analysis of cultivated melon groups (Cucumis melo L.) using random amplified polymorphic DNA and simple sequence repeat markers

Random amplified polymorphic DNA (RAPD) and simple sequence repeat (SSR) markers were used to characterize genetic relationships among 46 accessions in two C. melo L. subsp. melo (Cantalupensis, Inodorus) and subsp.agrestis (Conomon, and Flexuosus) groups. Genetic distance (GD) estimates were made among and between accessions in four melon market classes [Galia, Ogen, Charentais, and Shipper (European and U.S. types)] of Cantalupensis, one market class of Inodorus (Cassaba and Honey Dew), one accession of Conomon, and one accession of Flexuosus by employing three GD estimators; simple matching coefficient, Jaccard's coefficient, and Nei's distance-D. Differences detected among 135 RAPD bands and 54 SSR bands (products of 17 SSR primers) were used to calculate GD. Band polymorphisms observed with 21 RAPD primers and 7 SSR primers were important (p =0.01) in the detection of genetic differences. Estimators of GD were highly correlated (p 0.0001; r_s = 0.64 to0.99) when comparisons were made between estimation methods within a particular marker system. Lower correlations (r_s = 0.17 to 0.40) were detected (P > 0.001) between marker systems using any one estimator. The GD of the Conomon and Flexuosus accessions was significantly different (p> 0.001)from the mean GD of all the market classes examined. The mean GD (Jaccard's coefficient) among accessions of Ogen, Galia, Cassaba, Charentais, European shipper, and U.S. shipper groups was 0.11 ± 0.04, 0.33± 0.09, 0.21 ± 0.04, 0.26 ± 0.10, 0.17± 0.05 and 0.22 ± 0.08, respectively. Market classes were distinct (p > 0.001), such that GDs between Galia and other accessions were the largest(mean GD 0.34 to 0.35), and GDs between Ogen and other accessions were the smallest (mean GD 0.29 to 0.30). Contrasts between the U.S. shipper cultivar Top Mark and accessions within any market class was relatively large (mean GD = 0.42 ± 0.06). Empirical estimations of variances associated with each marker type in the accessions examined indicated that, per band, lower coefficients of variation can be attained in the estimation of GD when using RAPDs compared to SSRs. Nevertheless, the genetic relationships identified using these markers were generally similar. The disparity between the analyses of the two markers made may be related to the amount of genome coverage which is characteristic of a particular marker system and/or its efficiency in sampling variation in a population. Results of RAPD marker analysis suggest that 80 marker bands were adequate for assessing the genetic variation present in the accessions examined. This revised version was published online in July 2006 with corrections to the Cover Date.     

利用SRAP与SSR标记分析不同类型甜菜的遗传多样性

为选育优质甜菜新品种, 指导种质资源引进和利用, 为进行分子标记辅助选择育种提供科学依据, 采用SRAP和SSR两种分子标记方法相结合, 对甜菜单胚雄性不育系及保持系等49份材料进行遗传多样性分析。利用4个表型差异显著的甜菜品系对SRAP的64对引物组合及SSR的11对引物组合进行扩增, 分别筛选出有效引物组合11对和9对。SRAP的11对引物组合共产生199条扩增带, 其中有86条多态性带, 多态性带的比率平均为43.7%。SSR的9对引物共产生35条扩增带, 多态性比率为100%。全部材料的平均遗传距离为0.3860, 平均遗传相似系数为0.6795, 大约30%的材料遗传距离或遗传相似系数具显著或极显著差异。遗传相似系数平均值比较, 多胚四倍体品系0.7264＞单胚杂交组合0.7243＞国外品种0.7060＞多胚二倍体品系0.6908＞单胚品系0.6837。在遗传距离0.20处, 将49个甜菜材料划分为A、B、C、D 4个类群, D类群又分为4个亚类, 较好地显示了甜菜材料丰富的遗传多样性。表明不同甜菜品种具有相当高的异质性, 国外与国内材料的遗传基础存在一定差异, 但生产应用的甜菜品种间存在亲缘关系较近、遗传基础较窄的倾向。