AFLP analysis of Cynodon dactylon (L.) Pers. var. dactylon genetic variation.

Cynodon dactylon (L.) Pers. var. dactylon (common bermudagrass) is geographically widely distributed between about lat 45 degrees N and lat 45 degrees S, penetrating to about lat 53 degrees N in Europe. The extensive variation of morphological and adaptive characteristics of the taxon is substantially documented, but information is lacking on DNA molecular variation in geographically disparate forms. Accordingly, this study was conducted to assess molecular genetic variation and genetic relatedness among 28 C. dactylon var. dactylon accessions originating from 11 countries on 4 continents (Africa, Asia, Australia, and Europe). A fluorescence-labeled amplified fragment length polymorphism (AFLP) DNA profiling method was used to detect the genetic diversity and relatedness. On the basis of 443 polymorphic AFLP fragments from 8 primer combinations, the accessions were grouped into clusters and subclusters associating with their geographic origins. Genetic similarity coefficients (SC) for the 28 accessions ranged from 0.53 to 0.98. Accessions originating from Africa, Australia, Asia, and Europe formed major groupings as indicated by cluster and principal coordinate analysis. Accessions from Australia and Asia, though separately clustered, were relatively closely related and most distantly related to accessions of European origin. African accessions formed two distant clusters and had the greatest variation in genetic relatedness relative to accessions from other geographic regions. Sampling the full extent of genetic variation in C. dactylon var. dactylon would require extensive germplasm collection in the major geographic regions of its distributional range.     

野生狗牙根种质遗传多样性的SRAP研究

采用SRAP分子标记技术, 对采自中国四川、重庆、贵州、西藏四省区的32份野生狗牙根(Cynodon dactylon)材料进行遗传多样性分析, 获得下述结果：(1)用14对引物组合共得到132条多态性条带, 平均每对引物扩增出9.4条多态带, 多态性位点百分率为79.8%, 材料间的遗传相似系数范围在0.591到0.957之间, 平均GS值为0.759, 这些结果说明, 供试野生狗牙根具有较为丰富的遗传多样性; (2)对所有材料进行聚类分析, 可聚为4类, 大部分来自相同或相似生态地理环境的材料聚为一类, 表明供试材料的聚类和其生态地理环境间有一定的相关性; (3)基于Shannon多样性指数估算了6个狗牙根生态地理类群内和类群间的遗传分化, 发现类群内遗传变异占总变异的65.56%, 而类群间遗传变异占总变异的34.44%; (4)对各生态地理类群基于Nei氏无偏估计的遗传一致度的聚类分析表明, 各生态地理类群间的遗传分化与其所处的生态地理环境具有一定的相关性。     

湖南四种尾矿环境下的狗牙根遗传多样性的RAPD分析

选择湖南4种有代表性的有色金属尾矿地所生长的野生狗牙根作为研究对象,并以正常生境下的野生狗牙根作为对照,采用RAPD技术分析了这些特别环境下的狗牙根的遗传多样性。17条10个核苷酸长的引物用于PCR扩增,共得到432条DNA带。检测位点160个,其中多态位点134个,占83.75%。各个类群间的Jaccard相似系数平均为0.4475±0.0806,遗传距离平均为0.3825±0.0712。结果显示生长于这些不同尾矿环境下的狗牙根种群在遗传本质上均产生了明显的分化,完全可以根据其尾矿基质的不同而划分为不同的生态型或品种,甚至可能处理为新的变种。狗牙根种群具有丰富的遗传多样性,作为先锋植物可为尾矿的复垦提供更多的品种选择。     

狗牙根与非洲狗牙根形态鉴别性状的筛选

对10个狗牙根〔Cynodon dactylon(L.)Pers.〕和2个非洲狗牙根(C.transvaalensis Burtt-Davy)种源的13个外部形态性状进行了实地观测和统计分析。结果表明,与狗牙根相比,非洲狗牙根的叶片宽度较窄,穗支数较少,小穗较长,节间直径较细,穗支长度较短,差异显著;非洲狗牙根的叶片、葡匐茎及柱头的颜色与狗牙根有明显不同,因此,叶片宽度、节间直径、穗支数、穗支长度、小穗长度、叶片色泽、匍匐茎色泽和柱头色泽8个外部形态性状可作为鉴别这2个种的形态学指标。     

基于SRAP和TRAP标记的河北野生狗牙根种质遗传多样性分析

以SRAP和TRAP 2种标记技术对36份狗牙根材料的遗传多样性及亲缘关系进行了分析,其中包含34份河北省野生狗牙根种质资源。分别由238对SRAP和85对TRAP引物组合中筛选获得具有多态性的SRAP和TRAP引物组合各10对,PCR扩增总条带分别为186和161条,多态性条带156和132条,平均每对引物扩增出多态性条带各15.6和13.2条,多态性位点比率分别为83.4%和81.0%。2种标记合并进行聚类分析,所有供试的36份狗牙根材料遗传相似系数GS=0.519~0.983,平均为0.7。当GS=0.68时,可将36份供试材料分为4个类群。本研究结果表明河北野生狗牙根种质资源存在较丰富的遗传多样性,可为种质资源保护和选育优良狗牙根新品种提供科学依据。     

Elucidating polyploidization of bermudagrasses as assessed by organelle and nuclear DNA markers

Clarification of relationships among ploidy series of Cynodon accessions could be beneficial to bermudagrass breeding programs, and would enhance our understanding of the evolutionary biology of this warm season grass species. This study was initiated to elucidate polyploidization among Cynodon accessions with different ploidy series collected from Turkey based on chloroplast and nuclear DNA. Forty Cynodon accessions including 7 diploids, 3 triploids, 10 tetraploids, 11 pentaploids, and 9 hexaploids were analyzed using chloroplast DNA restriction fragment-length polymorphism (cpDNA RFLP), chloroplast DNA simple sequence repeat (cpDNA SSR), and nuclear DNA markers based on neighbor-joining (NJ) and principle component analyses (PCA). All three-marker systems with two statistical algorithms clustered the diploids apart from the other ploidy levels. Assuming autopolyploidy, spontaneous polyploidization followed by rapid diversification among the higher ploidy levels than the diploids is likely in Cynodon's evolution. Few tetraploid and hexaploid accessions were clustered with or closely to the group of diploids, supporting the hypothesis above. Eleven haplotypes as estimated by cpDNA RFLP and SSR markers were detected. This study indicated that the diploids had different organelle genome from the rest of the ploidy series and provided valuable insight into relationships among ploidy series of Cynodon accessions based on cp and nuclear DNAs.     

Phylogeny of ten species of the genus Hordeum L. as revealed by AFLP markers and seed storage protein electrophoresis

The phylogenetic relationships of 60 accessions representing ten species of the genus Hordeum were investigated based on AFLP markers and seed storage protein SDS-PAGE electrophoresis. A total of 339 AFLP polymorphic markers were scored as a result of fingerprinting the studied taxa using seven AFLP primer combinations, whereas 46 polymorphic protein bands resulted from the water soluble and water non-soluble seed storage protein electrophoresis. The phylogenetic tree deduced from AFLP analysis is concordant in a large extent with that deduced from seed storage protein electrophoresis. The studied taxa were clustered according to their genome type into two main groups representing the Old and New World’s species. Inside each group the species were clustered according to their genome type. Highly significant cophenetic correlation coefficient was obtained between both AFLP (0.96) and seed storage protein (0.89) indicating the reliability of the results.     

HYBRIDIZATION STUDIES WITH SPECIES OF CYNODON FROM EAST AFRICA AND MALAGASY

A collection of 198 accessions of Cynodon from East Africa and Malagasy was grown in uniform nurseries at Stillwater, Okla., and classified morphologically, studied cytologically, and tested for crossability in hybridization studies. The following taxa were recognized: C. arcuatus, C. plectostachyus, C. aethiopicus, C. nlemfuensis var. nlemfuensis, C. nlemfuensis var. robustas, C. dactylon var. aridus, C. dactylon var. coursii, and C. dactylon var. dactylon. The magnitude of barriers to gene flow was estimated by ease of hybridization and by sterility of the hybrids obtained. C. arcuatus and C. plectostachyus are well isolated from all other species in the genus. The others are sufficiently related that they may be intercrossed. The barriers to gene flow vary demonstrably in strength between different combinations of taxa. The magnitude of a barrier required to establish species limits is, perhaps, more a matter of subjective opinion than a scientific conclusion. It was concluded that no completely satisfactory classification of the genus is possible. The system adopted was a compromise between classical morphological concepts of species, the biological concept of species, and pragmatic utility.     

Phylogenetic analysis reveals multiple introductions of Cynodon species in Australia

The distinction between native and introduced flora within isolated land masses presents unique challenges. The geological and colonisation history of Australia, the world's largest island, makes it a valuable system for studying species endemism, introduction, and phylogeny. Using this strategy we investigated Australian cosmopolitan grasses belonging to the genus Cynodon. While it is believed that seven species of Cynodon are present in Australia, no genetic analyses have investigated the origin, diversity and phylogenetic history of Cynodon within Australia. To address this gap, 147 samples (92 from across Australia and 55 representing global distribution) were sequenced for a total of 3336bp of chloroplast DNA spanning six genes. Data showed the presence of at least six putatively introduced Cynodon species (C. transvaalensis, C. incompletus, C. hirsutus, C. radiatus, C. plectostachyus and C. dactylon) in Australia and suggested multiple recent introductions. C. plectostachyus, a species often confused with C. nlemfuensis, was not previously considered to be present in Australia. Most significantly, we identified two common haplotypes that formed a monophyletic clade diverging from previously identified Cynodon species. We hypothesise that these two haplotypes may represent a previously undescribed species of Cynodon. We provide further evidence that two Australian native species, Brachyachne tenella and B. convergens belong in the genus Cynodon and, therefore, argue for the taxonomic revision of the genus Cynodon.     

AFLP variation in 25 Avena species

Current molecular characterization of ex situ plant germplasm has placed more emphasis on cultivated gene pools and less on exotic gene pools representing wild relative species. This study attempted to characterize a selected set of germplasm accessions representing various Avena species with the hope to establish a reference set of exotic oat germplasm for oat breeding and research. The amplified fragment length polymorphism (AFLP) technique was applied to screen 163 accessions of 25 Avena species with diverse geographic origins. For each accession, 413 AFLP polymorphic bands detected by five AFLP primer pairs were scored. The frequencies of polymorphic bands ranged from 0.006 to 0.994 and averaged 0.468. Analysis of molecular variance revealed 59.5% of the total AFLP variation resided among 25 oat species, 45.9% among six assessed sections of the genus, 36.1% among three existing ploidy levels, and 50.8% among eight defined genome types. All the species were clustered together according to their ploidy levels. The C genome diploids appeared to be the most distinct, followed by the Ac genome diploid A. canariensis. The Ac genome seemed to be the oldest in all the A genomes, followed by the As, Al and Ad genomes. The AC genome tetraploids were more related to the ACD genome hexaploids than the AB genome tetraploids. Analysis of AFLP similarity suggested that the AC genome tetraploid A. maroccana was likely derived from the Cp genome diploid A. eriantha and the As genome diploid A. wiestii, and might be the progenitor of the ACD genome hexaploids. These AFLP patterns are significant for our understanding of the evolutionary pathways of Avena species and genomes, for establishing reference sets of exotic oat germplasm, and for exploring new exotic sources of genes for oat improvement.     

中国狗牙根(Cynodon dactylon)优良选系的RAPD分析

对采自中国不同地区的6份狗牙根[Cynodon dactylon (L.) Pers.]优良选系进行了RAPD分子标记实验.选用15个引物共扩增出438条带,平均每个引物扩增出29条带,其中多态性带415条,多态性位点百分率达到59.02%～75.49%.各选系间遗传相似性系数差异较大(0.408～0.672),说明各选系间在DNA水平上存在着丰富的遗传多样性,与新品种C106(爬地青)相比,各选系还存在一定的改良空间.实验结果也表明,RAPD分子标记可成功地用于中国狗牙根优良选系遗传多样性的研究及品种鉴定.     

Leaf blade structure and C4 acid decarboxylation enzymes in xCynochloris spp. (Poaceae), intergeneric hybrids between species of different C4 type

PRENDERGAST, H. D. V., STONE, N. E. & HATTERSLEY, P. W., 1988. Leaf blade structure and C₄ acid decarboxylation enzymes in x Cynochloris spp. (Poaceae), intergeneric hybrids between species of different C₄ type. x Cynochloris macivorii Clifford and Everist and x C. reynoldensis B. K. Simon (Poaceae) are intergeneric C₄ hybrids between Cynodon dactylon (L.) Pers., and NAD-malic enzyme (NAD-ME) species, and two different PEP carboxykinase (PCK) species of Chloris . Parental species of each hybrid species have the 'classical' leaf blade structure of their respective C₄ acid decarboxylation types. The outline of the photosynthetic carbon reduction (PCR or Kranz) bundle sheath and the position of the PCR cell chloroplasts in x Cynochloris are intermediate between those of the parental species, C. macivorii being more like Cynodon dactylon and x C. reynoldensis more like Chloris spp. The PCR chloroplast shape in x C. macivorii and x C. reynoldensis is like that of Cynodon dactylon and Chloris spp., respectively. Differences between the hybrids in their enzyme activities complement these structural differences: x C. macivorii has more NAD—ME and less PCK activity than x C. reynoldensis , although in both species PCK activity is the greater. Both hybrids, however, have a suberized lamella in PCR cell walls as do Chloris spp. The close taxonomic relationship between Cynodon and Chloris make these genera especially suitable for reciprocal crossing experiments aimed at increasing understanding of the genetic relationships between subtypes of the C₄ photosynthetic pathway.     

Molecular phylogeny and DNA amplification fingerprinting of Petunia taxa

The relationship of five species of Petunia and ten cultivars of the cultivated petunia, Petunia x hybrida, were investigated using DNA-amplification fingerprinting (DAF). Reproducible banding profiles were obtained from P. parodii and P. axillaris DNA from different seed sources. In contrast, other petunias such as P. inflata, P. violacea and P. integrifolia produced variable fingerprints when different plants were examined. However, representative profiles of the variable Petunia taxa were obtained by bulking the leaf tissue from ten different individual plants. Each of ten octamer primers revealed polymorphic loci between taxa. Among a total of 201 bands produced, 146 (73%) loci were polymorphic and distinguished all species and cultivars. Phenetic and cluster analysis using DAF markers separated P. axillaris from P. parodii and distinguished between the violet-flowered species, P. inflata, P. violacea, and P. integrifolia. P. parodii grouped together with the monophyletic set of the ten cultivars of P. x hybrida examined, indicating that it had made a major contribution to the development of these cultivars. Cultivars were distributed within the dendograms by flower color. The results demonstrated the utility of DAF in establishing relationships among closely related species and cultivars of Petunia.     

短柄枹种群遗传多样性的ISSR分析

利用ISSR分子标记方法对分布在浙江省境内的7个短柄枹种群的遗传多样性和遗传分化进行了分析。从100个引物中筛选出12个用于正式扩增的ISSR引物,在7个种群140个个体中共检测到132个位点,其中多态位点118个,多态位点百分率(P)为89.39%,各种群P平均为58.87%。短柄枹总的Shannon信息指数(I)为0.493 3、Nei指数(h)为0.334 7,各种群I平均为0.336 2、h平均为0.229 1。P、I、h均显示云峰种群最高,天台山种群最低。AMOVA分子差异分析表明,67.97%的变异存在于种群内,32.03%的变异存在于种群间,种群间的基因分化系数(G_ST)为0.315 4。短柄枹种群间的基因流为(N_m)为1.085 3。7个种群的平均遗传距离为0.173 9。利用UPGMA法对7个种群进行聚类,结果显示天台山和雪窦山种群聚成一类,其它5个种群聚成另一类。     

Estimation of genetic variation in Cynodon dactylon accessions using the ISSR technique

Genetic variation in 55 accessions of Cynodon dactylon was estimated using inter-simple sequence repeat (ISSR) markers. The plant materials used in this study originated from 17 countries. A total of 236 ISSR fragments were generated with 14 primers. Fragment sizes ranged from 200 to 3000 bp. All scorable bands were polymorphic in nature and none of the primers used produced monomorphic bands, indicating a high level of genetic variation in this grass. The accessions were found to be clustered into eight major groups through the unweighted pair-group method with arithmetic averages. Genetic similarity coefficients (GSC) among the 55 accessions ranged from 0.52 to 0.95. The results clearly indicate that a high level of variation exists in Cynodon accessions. This study shows that the ISSR technique is a reliable tool for differentiating Cynodon accessions and for determining the genetic relationships among them.     

Relationships among <Emphasis Type="Italic">Leymus</Emphasis> species assessed by RAPD markers

The DNA genetic diversity of 40 accessions of genus Leymus was analyzed by random amplified polymorphic DNA (RAPD) markers. A total of 352 products were amplified by 34 10-mer arbitrary primers, among which 337 products (95.74 %) were found to be polymorphic. 5–14 polymorphic bands were amplified by each polymorphic primer, with an average of 9.91 bands. The data of 352 RAPD bands were used to generate Jaccard’s similarity coefficients and to construct a dendrogram by means of UPGMA. Great genetic diversity in genus Leymus was observed, the genetic diversity among the different species more abundant than that of the different accessions, and the different accessions in a species or the species from the same areas were clustered together.     

Modified AFLP technique for rapid genetic characterization in plants

The standard amplified fragment-length polymorphism (AFLP) technique was modified to develop a convenient and reliable technique for rapid genetic characterization of plants. Modifications included (i) using one restriction enzyme, one adapter molecule and primer, (ii) incorporating formamide to generate more intense and uniform bands and (iii) using agarose gel electrophoresis. Sea oats (Uniola paniculata L.), pickerel-weed (Pontederia cordata L.), Bermudagrass (Cynodon dactylon L.) and Penstemon heterophyllus Lindl. were used to determine the ability to generate adequate resolution power with both self- and cross-pollinated plant species including cultivars, ecotypes and individuals within populations. Reproducibility of bands was higher in all the AFLP experiments compared to random amplified polymorphic DNA (RAPD). Formamide with or without bovine serum albumin improved band intensities compared to dimethyl sulfoxide and the standard reaction mixture with no organic solvents. Comparison between RAPD and modified AFLP using sea-oats population samples proved that modified AFLP exhibits (i) a low number of faint bands with increased specificity of amplified bands, (ii) a significantly higher number of polymorphic loci per primer, (iii) less primer screening time, (iv) easy scoring associated with fewer faint bands and (v) greatly enhanced reproducibility. The technique described here can be applied with a high degree of accuracy for plant genetic characterization.     

用ISSR标记检测黄麻野生种与栽培种遗传多样性

利用ISSR标记对黄麻属((Corchorus)10个种27份材料的遗传多样性进行分析,25个ISSR引物共扩增出283条带,平均每个引物扩增出10．48条带,多态性条带比例(PPB)为92．85％;种间遗传相似系数在0．33～0．97之间。表现出丰富的遗传多样性．系统聚类结果显示,第I、Ⅲ类群均为黄麻属8个种11份原始野生种,种间存在丰富的遗传多样性;第Ⅱ类群为黄麻两个栽培种及其近缘野生种,共16份材料。种遗传相似性较高;基因组聚类结果与经典分类相符．利用分子标记技术研究初步可以认为。荨麻叶种为最原始的黄麻野生种之一;三室种21C为三室种的一个变种;甜麻为一个尚待定性的野生种．同组材料的RAPD和ISSR分析结果,具有较高的相似性和可信度．     

Genetic diversity in tetraploid switchgrass revealed by AFLP marker polymorphisms

Switchgrass (Panicum virgatum) is a perennial warm-season grass native to North America that has been identified as a dedicated cellulosic biofuel crop. We quantified genetic diversity in tetraploid switchgrass germplasm collected at Oklahoma State University and characterized genetic relatedness among the collections from distinct regions. Fifty-six tetraploid accessions, including seven upland and 49 lowland genotypes from throughout the US, were examined. The amplified fragment length polymorphism (AFLP) procedure was utilized to generate DNA profiling patterns that were scored visually. Sixteen selective AFLP primer combinations were used to amplify 452 polymorphic bands. The accessions' genetic similarity coefficients, UPGMA (unweighted pair-group method with arithmetic averaging) cluster analysis and principle coordinate analysis, were performed. The upland and lowland accessions clustered according to ecotypes, with one exception (TN104). Genetic similarity coefficients among the accessions ranged from 0.73 to 0.95. Analysis of molecular variance (AMOVA) was performed, showing significant differences between the upland and lowland genotypes. The trnL marker confirmed that TN104 was a lowland genotype, but the trnL marker identification of upland and lowland genotypes was not consistent with the AFLP analysis in two germplasms (Miami and AR4).