Isozyme, ISSR and RAPD profiling of genotypes in marvel grass (Dichanthium annulatum)

Genetic analysis of 30 accessions of marvel grass (Dichanthium annulatum Forsk.), a tropical range grass collected from grasslands and open fields of drier regions, was carried out with the objectives of identifying unique materials that could be used in developing the core germplasm for such regions as well as to explore gene (s) for drought tolerance. Five inter-simple sequence repeat (ISSR) primers [(CA)4, (AGAC), (GACA) 4; 27 random amplified polymorphic DNA (RAPD) and four enzyme systems were employed in the present study. In total, ISSR yielded 61 (52 polymorphic), RAPD 269 (253 polymorphic) and enzyme 55 isozymes (44 polymorphic) bands. The average polymorphic information content (PIC) and marker index (MI) across all polymorphic bands of 3 markers systems ranged from 0.419 to 0.480 and 4.34 to 5.25 respectively Dendrogram analysis revealed three main clusters with all three markers. Four enzymes namely esterase (EST), polyphenoloxidase (PPO), peroxidase (PRX) and superoxide dismutase (SOD) revealed 55 alleles from a total of 16 enzyme-coding loci. Of these, 14 loci and 44 alleles were polymorphic. The mean number of alleles per locus was 3.43. Mean heterozygosity observed among the polymorphic loci ranged from 0.406 (SOD) to 0.836 (EST) and accession wise from 0.679 (1G3108) to 0.743 (IGKMD-10). Though there was intermixing of few accessions of one agro-climatic region to another largely groupings of accessions were with their regions of collections. Bootstrap analysis at 1000 iterations also showed large numbers of nodes (11 to 17) having strong clustering (> 50 bootstrap values) in all three marker systems. The accessions of the arid and drier regions forming one cluster are assigned as distinct core collection of Dichanthium and can be targeted for isolation of gene (s) for drought tolerance. Variations in isozyme allele numbers and high PIC (0.48) and MI (4.98) as observed with ISSR markers indicated their usefulness for germplasm characterization.     

11个猕猴桃品种间的遗传多样性分析

利用随机扩增多态性DNA标记(RAPD)技术对采自四川省猕猴桃科研基地的11个猕猴桃品种进行了遗传多样性分析.通过引物筛选,从15个RAPD引物中扩增出135条带,其中100条为多态带,占74.07%.UPGMA聚类分析结果揭示了各品种间的亲缘关系.RAPD标记说明猕猴桃品种间遗传距离与地理分布有一定关系,地理位置较近的材料能聚在一起.分子标记可用于猕猴桃种质资源的分类、鉴定及良种选育.图2表2参15     

Use of EST database markers from M. truncatula in the transferability to other forage legumes

In general tropical forage legumes lack microsatellites or simple sequence repeat (SSR) markers. Development of genic SSR markers from expressed sequence tagged (EST) database is an alternate and efficient approach to generate the standard DNA markers for genome analysis of such crop species. In the present paper a total of 816 EST-SSRs containing perfect repeats of mono (33.5%), di (14.7%), tri (39.3%), tetra (2.7%), penta (0.7%) and hexa (0.4%) nucleotides were identified from 1,87,763 ESTs of Medicago truncatula. Along with, 70 (8.5%) SSRs of a compound type were also observed. Seven primer pairs of tri repeats were tested for cross transferability in 19 accessions of forage legumes comprising 11 genera. At two different annealing temperatures (55 and 60 degreesC) all primer pairs except AJ410087 reacted with many accessions of forage legumes. Atotal of 51 alleles were detected with six M. truncatula EST-SSRs primer-pairs against DNAfrom 19 accessions representing 11 genera where number of alleles ranged from 2 to 13. The cross-transferability of these EST-SSRs was 40.6% at 55 degreesC and 32.3% at 60 degreesC annealing temperature. 24 alleles of the total 50 (48%) at 55 degreesC and 27 of 51 (53%) at 60 degreesC were polymorphic among the accessions. These 27 polymorphic amplicons identified could be used as DNA markers. This study demonstrates the developed SSR markers from M. truncatula ESTs as a valuable genetic markers and also proposes the possibility of transferring these markers between species of different genera of the legumes of forage importance. It was evident from the results obtained with a set of Desmanthus virgatus accessions where SequentialAgglomerative Hierarchical and Nested (SAHN) cluster analysis based on Dice similarity and Unweighted Pair Group Method with Arithmetic mean Algorithm (UPGMA) revealed significant variability (24 to 74%) among the accessions. High bootstrap values (>30) supported the nodes generated by dendrogram analysis of accessions.     

Identification of Tsuga Germplasm by Morphological Characters and RAPD Markers

Germplasm collection is important to preserve and maximize genetic diversity for germplasm conservation. Tsuga dumosa （ D. Don） Eichler in Engler ＆ Prantl. and T. chinensis var. forrestii （Downie） Silba germplasm was collected from three localities in China： Mt. Yulong, Wenfeng Temple and Mt. Dishiergu, Yunnan Province. Accessions were identified based on morphological characters and RAPD markers. The shapes of the apices and margins of needles were examined, and the length and width of needles, cones and seeds from accessions of mature plants were used to compare the morphological differences and to identify the germplasm. Molecular markers generated by randomly amplified polymorphic DNA （RAPD） were also used to characterize the taxa. Although the clustering based on RAPD markers was inconsistent with the morphological characters of the needles, based on the overall morphological characters and on RAPD markers, the accessions from Mt. Yulong and Wenfeng Temple were identified as T. chinertsis var. forrestii, and those from Mt. Dishiergu identified as T. dumosa. Taxonomic identification of the accessions was made based on morphology and by RAPD markers concurred. The results indicate that the shapes of the apices and margins of needles particularly from young plants could not be used as a possible key to identify T. dumosa and T. chinertsis var. forrestii. Fig 6, Tab 3, Ref24     

用SSR标记检测杂交籼稻三系亲本的遗传差异

随机选用分布于水稻(Oryza sativa L.)12条染色体上的110对引物,共筛选出26对具有多态性的SSR引物,对24份杂交籼稻三系亲本材料进行遗传多样性和亲缘关系分析.全部24份亲本中共检测出116对等位基因,每个SSR引物可检测到的等位基因数目为2～7个不等,平均每对引物可以检测到4.4个等位基因.多态信息量(PIC)的变动范围为0.068 9～0.803 6,平均PIC值为0.603 4;期望杂合度(He)的变动范围为0.081 6～0.829 8,平均值为0.616 9;多样性指数(1)变幅为0.173 2～1.791 3,平均为1.160 0.对供试亲本材料根据遗传距离进行UPGMA聚类分析,结果表明,SSR微卫星标记可以将24份亲本材料分为3个大群,各组亲本间的遗传差异较大.通过分子标记辅助育种,可以有效地鉴定供试亲本材料之间的亲缘关系,分析预测新种质材料的遗传差异,可作为选育强优势组合杂交水稻的重要参考指标之一.图2表3参24     

Identification of species-specific RAPD markers in genus Cenchrus

Cenchrus is an important component of major grass cover of world. Similar to the other major tropical grasses most of the species in genus Cenchrus are also apomictic in nature hence correct and precise identification of accessions and species are problematic and dubious. In the present study 187 decamer oligonucleotide primers were tested for PCR-based DNA amplification of six prominent species of genus Cenchrus. Of these, 32 potential repetitive and polymorphic primers were tested for identification of species-specific markers for C. ciliaris, C. setigerus, C. pennisetiformis, C. prieurri, C. biflorus and C. myosuroides. These primers yielded 51 unique RAPD markers either specific to a species (37) or shared by two or more species (14). Maximum markers were shared between C. ciliaris and C. setigerus confirming theirmore closeness to each other Primers like OPF09, OPF11, OPR15, OPAJ11, OPQ10 and OPAK20 generated strong intense bands can be used on priority in identifying the species from their natural habitat for the development of species-specific core germplasm. Due to apomictic nature this is the prime method of developing cultivars, as morphological characters are largely unable to distinguish them. The level of variation observed clearly suggest RAPD as an appropriate marker for genetic studies and in identifying the lines with species-specific markers for Cenchrus germplasm management activity and also maintaining identity and purity for proprietary reasons.     

杉木地理种源遗传变异的RAPD分析

应用随机扩增多态性DNA技术对我国12个有代表性的杉木地理种源遗传多样性进行分析，从80个随要引物中筛选出20个进行扩增，共扩增出149个DNA片段，其中具多态性的片段有115个，占77．18％％，表明极木地理种源间具有丰富的DNA序列多态性、平均每个引物提供7．45个RAPD标记的信息量，扩增出的DNA片段大小一般为150到2000bp范围之间，12个杉木地理种源间遗传距离变幅为0．193－0．4467，聚类结果表明，在0．2664处，广东信宜，广西梧州，湖南会同，湖南江华，贵州锦屏，江西全南聚为一类，福建沙县，浙江开化，湖北咸宁，安徽休宁聚为一类，四川雅安，陕西南郑各为一类，图2表2参15     

柚类资源及其近缘种SSR标记的分子评价

采用31对SSR引物对122份柚类种质资源及其近缘种的遗传多样性进行了研究.31对SSR引物可以检测到335个等位基因变异,平均每个位点可检测到9.85个等位基因.每个位点多态性信息指数(PIC)平均值为0.7085.从这些引物中有效地筛选出21对高信息量SSR引物,与31对引物在122份柚类资源间遗传关系达到极显著相关(r=0.99104).柚类资源及其近缘种等位基因平均数(A)、平均杂合位点百分比(P)、SSR表型杂合度(H0)分别为2.8、83.54%、0.524.用UPGMA方法将122份研究材料分成7个组群,110个柚类品种在相似系数0.712时可细分成18个亚组,分类结果有利于今后有目的地利用这些丰富育种材料的遗传背景.图3表2参21     

山西高原油松种群遗传多样性

用酸性聚丙烯酰胺凝胶电泳（A-PAGE）技术，分析了山西高原9个油松种群在醇溶蛋白水平上的遗传多样性。135份材料共分离出23条带，其中3条为共有带，多态性高达86．95％。全部材料共出现53种带型，9个不同油松种群的带型有差异，同一种群不同个体的带型也有所不同，说明山西高原的油松在遗传上已产生一定程度分化，在醇溶蛋白水平上呈现出遗传多态性。从供试材料的带型计算出油松遗传分化系数为0．1547。即在种群间的变异占总变异的15．47％，种群内变异为84．53％，大部分的遗传变异存在于种群内，但种群间的分化程度在松属树种中也属于较高水平。根据23个多态位点计算遗传相似系数和遗传距离，进行聚类分析，将山西高原9个油松种群聚为3个类群。     

中国茸鹿品种(品系)的随机扩增多态DNA(RAPD)研究

采用随机扩增多态ＤＮＡ技术研究了５个茸鹿品种（品系）７５个个体的遗传变异关系。在所使用的４１个引物中，有４０个引物扩增出多态谱带，共检测到３９５条扩增片段，其中２５９条（６５．６％）出现变异，用Ｓｈａｋｎｎｏｎ指数计算了５个品种（品系）的遗传变异及其遗传变异在群体内和群体间的分布。利用Ｎｅｉ氏片段共享度计算了７５个个体间的遗传距离，用ＵＰＧＭＡ聚类法构建了７５个个体的系统发育树状图，反映５个茸鹿品     

利用RAPD标记分析卧龙自然保护区不同海拔沙棘种群的遗传变异

从1800m到3400m五个海拔连续取样,用RAPD分子标记研究了卧龙自然保护区中国沙棘种群的遗传结构和遗传变异.用11条寡核苷酸引物,扩增得到151个重复性好的位点,其中143个多态位点,多态率达94.7%.在5个沙棘种群中,总遗传多样性值(HT)为0.289,B种群内的遗传多样性值为0.315,这完全符合沙棘这种多年生、远交的木本植物高遗传变异的特性.5个种群内遗传多样性随海拔升高呈低—高—低变异趋势,在2200m海拔处的B种群遗传多样性达最大值0.315,3400m海拔处的E种群则表现最小,仅0.098.5个种群间的遗传分化值GST=0.406,也即是说有40.6%的遗传变异存在于种群间,59.4%存在种群内.1800m海拔处的A种群与其他种群的明显分离是造成种群间遗传分化大的原因.UPGMA聚类图和PCoA散点图分别进一步确证了5个种群间关系和所有个体间的关系.最后,经过Mantel检测,遗传距离与海拔表现了明显的相关性(r=0.646,P=0.011).     

RAPD方法鉴定油菜“杂油59”种子纯度的研究

本文采用ＲＡＰＤ分子标记技术对我国育成的油菜新品种“杂油５９”的亲本和Ｆ１代种子纯度进行了技术鉴定。用４０个１０ｍｅｒ的随机引物对“杂油５９”的２个亲本“垦Ｃ８”和“陕３Ａ”进行了ＲＡＰＤ分析，共出现２９０条带，分布于３５３０～２２０ｂｐ之间，大部分条带在双亲之间是相同的。引物Ｏｐａ－Ｏ６，Ｏｐｃ－１２，Ｏｐｃ－２０，Ｏｐｋ－０３，ｏｐｋ－１３，Ｏｐｊ－１２出现差异扩增带，经实验确认Ｏｐｋ－０３的     

Genetic Diversity in a Threatened Wetland Species, Helonias bullata (Liliaceae)

Genetic variation was examined in Helonias bullata , a threatened perennial plant species that occurs in isolated wetland habitats. Fifteen populations representing the species' geographic range were sampled. Genetic diversity was low for the species ( H _es = 0.053) as well as within populations ( H _ep = 0.029). Of the 33 allozyme loci examined, 11 (33%) were polymorphic, while on average only 12.8% (4) of the loci were polymorphic within populations. The number of alleles per polymorphic locus was 2.36 for the species and averaged 2.09 across populations. For every genetic parameter calculated, variation in H. bullata was lower than that typically found for narrowly distributed plant species. The lowest levels of genetic diversity were found in northern areas that were colonized following the last glacial epoch. The number of genotypes detected per population ranged from three to 21, with a mean of 13 for this clonally reproducing species. We found a relatively high proportion of total genetic diversity (30.6%) among populations and a significant correlation (p < 0.002) between genetic distance and geographic distance. Genetic drift phenomena appear to play a major role in the population genetics of this species. Anomalously, several populations that appeared most limited in size and vigor were genetically most variable, perhaps because they represent older, relictual populations. Life-history characteristics of H. bullata coupled with low levels of genetic diversity and the degradation and disappearance of wetlands threaten the existence of this species.     

中国对虾两个不同地理种群遗传结构的RAPD分析

采用ＲＡＰＤ技术对中国对虾中国黄渤海沿岸种群（ＨＢ）和朝鲜半岛西海岸种群（ＰＫ）的遗传多样性进行检测。２０个随机引物（ＯＰＤ系列）在二个种群中都扩增出１４８条ＤＮＡ片段，其中多态性片段数分别为５４条和６５条，多态性片段的比例分别为３６．４９％和４３．９２％，种群的平均杂合度分别为０．１９８１和０．２３７５。实验表明，中国对虾朝鲜半岛西海岸种群的遗传多样性要比中国黄渤海沿岸种群高，但二个种群都处于较     

Genetic studies in marine fishes

Nine polymorphic loci were found among 42 presumptive protein-coding gene loci surveyed among 474 red drum (Sciaenops ocellatus) sampled in 1987 from 13 nearshore and 1 offshore localities from the Atlantic coast of the southeastern USA and the northern Gulf of Mexico. The mean number of alleles over the polymorphic loci was 3.8, and the average heterozygosity over all loci examined was estimated as 0.047. These data indicate that red drum have normal levels of genetic variability. Wright'sF1 _ST values (the standardized variance of allele frequencies between samples) over all polymorphic loci ranged from 0.009 to 0.027 (meanF1 _ST =0.019), and estimates of the effective number of migrants (N _e m) per generation using Wright's island model ranged from 9.0 to 27.5. High levels of gene flow among the red drum samples were also indicated by Slatkin's qualitative analysis using conditional average allele frequencies. Nei's estimates of genetic distance between pairs of samples ranged from 0.000 to 0.009, indicating a high degree of nuclear gene similarity among all samples. Highly significant heterogeneity in allele frequencies at the locus for adenosine deaminase was detected between red drum sampled from the Atlantic and those sampled from the Gulf and among red drum sampled from the Gulf.     

朱■的随机扩增多态DNA分析与种内亲缘关系研究

利用随机扩增多态ＤＮＡ技术对朱（Ｎｉｐｐｏｎｉａｎｉｐｐｏｎ）８个个体进行了随机扩增多态ＤＮＡ分析．用２０个１０ｂｐ的随机引物对每只朱的基因组ＤＮＡ进行扩增,共得到１６８个扩增片段,其中共有片段为１０２个．根据聚类分析所得到的树状图确定了８只朱的亲缘关系,这为进一步构建全部朱个体的谱系关系图打下了基础,有利于制定更有效的朱保护计划     

朱鹮的随机扩增多态DNA分析与种内亲缘关系研究

利用随机扩增多态DNA技术对朱鹮 ( Nipponia nippon ) 8个个体进行了随机扩增多态DNA分析. 用20个10bp的随机引物对每只朱鹮的基因组DNA进行扩增,共得到168个扩增片段,其中共有片段为102个. 根据聚类分析所得到的树状图确定了8只朱鹮的亲缘关系,这为进一步构建全部朱鹮个体的谱系关系图打下了基础,有利于制定更有效的朱鹮保护计划.     

RAPD markers suggest genotypic effects on forager specialization in a eusocial wasp

Genetic variability within insect societies may provide a mechanism for increasing behavioral diversity among workers, thereby augmenting colony efficiency or flexibility. In order to assess the possibility that division of labor has a genetic component in the eusocial wasp Polybia aequatorialis, I asked whether the genotypes of workers within colonies correlated with behavioral specialization. Workers specialized by foraging for one of the four materials (wood pulp, insect prey, nectar, or water) gathered by their colonies. I collected foragers on 2 days from each of three colonies and identified the material the foragers were carrying when collected. I produced random amplified polymorphic DNA (RAPD) markers from the genomic DNA of these foragers and estimated genotypic similarity of foragers based on sharing of variable RAPD marker bands. Contingency tests on 20 variable loci per colony showed statistically significant (P <0.05) biases in RAPD marker frequencies among forager types in the three colonies. Patterns of association of RAPD marker bands with specializations were constant in two colonies, but changed between collection days in one colony. RAPD marker biases suggest that division of labor among workers includes a genetic component in P. aequatorialis. Colony-level selection on variation in division of labor is a possible factor favoring the evolutionary maintenance of high genotypic variability (low relatedness) in epiponine wasp colonies and in other eusocial insects. Received: 18 July 1995/Accepted after revision: 1 October 1995     

Genetic mating system of the brown smoothhound shark (<Emphasis Type="Italic">Mustelus henlei</Emphasis>), including a literature review of multiple paternity in other elasmobranch species

Although an understanding of mating systems is thought to be an important component of long-term population management, these life history characteristics are poorly known in sharks. Here, we employ polymorphic microsatellite markers to test for the occurrence and prevalence of multiple paternity in a population of the brown smoothhound shark, Mustelus henlei. We analyzed litters from 14 females sampled from the Pacific coast of Baja California Sur. The minimum number of sires ranged from one to three with an average of 2.3 sires per litter. Regression analyses did not indicate a relationship between female body size and number of sires, or female body size and size of the litter. A review of the existing literature on genetic mating systems in sharks suggests that polyandry may be common and that reproductive behavior may have evolved from conflicting selection pressures between the sexes.     

Assessment of genetic variation in lucerne (Medicago sativa L.) using protease inhibitor activities and RAPD markers

The aim of the present study is to identify and characterize lucerne lines resistance to weevil infestation. After three years of field screening for resistance to weevil infestation, 13 lines of lucerne were selected to assess the genotypic variations for lucerne weevil (Hypera postica Gyll.) at biochemical and molecular levels. Total phenols varied from 0.15 to 0.91 mg g (DM) in these genotypes. The highest trypsin (11.11 unit mg(-1) protein) and chymotrypsin (93.0 unit mg(-1) protein) inhibitors activities were recorded in G-1-02 and B-4-03 lines respectively, whereas highest alpha-amylases inhibitor activity (14.2 unit mg(-1) protein) in C-6-01. Zymogram patterns for trypsin inhibitor activity showed quantitative variations among the lines. In total 262 DNA fragments were generated when 45 deca-mer random primers were employed. Genetic variation in terms of genetic distance ranged from 0.65 to 0.85. Sequential Agglomerative Hierarchical and Nested (SAHN) clustering using the Un-weighted Pair Group Method with Arithmetic mean (UPGMA) algorithm yielded two clusters (cluster I and II) which converged at 72% similarity level. Cluster I contained most of the lines having low level of weevil infestation. High bootstrap values (>40) indicated the significance of nodes embodied in these two clusters. However, SDS-PAGE analysis of the leaf proteins of these 13 lines showed no major variations except minor difference in the protein bands of molecular weights between 14 to 20 kD.