鸭茅种质资源遗传多样性的SRAP研究

利用SRAP(Sequence-related amplified polymorphism)标记对来自国内外的45份鸭茅（Dactylis glomerata L.）种质资源进行遗传多样性研究。21对引物扩增出438个条带,多态性条带为363条,多态性条带比率为82.08%,每对引物组合的多态性带数平均为17.29条。GS值范围在0.6248-0.9686间,平均GS值为0.7958,显示来源广泛的鸭茅种质资源间存在着丰富的遗传变异。聚类分析及主成分分析能将所有材料聚为4类,能较准确的反映材料的来源分布情况及供试材料的染色体倍性差异,表明鸭茅的遗传多样性与染色体倍性及地理分布密切相关。同时清楚的揭示出国产鸭茅品种遗传基础较为狭窄。本研究为育种和探讨鸭茅种质资源遗传变异奠定了较好的理论基础。     

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。     

鸭茅种质资源遗传多样性的ISSR研究

采用ISSR分子标记技术对来自国内及亚洲、欧洲、美洲9个国家共50份鸭茅品种(系)进行遗传多样性研究。12个引物共扩增出多态性带101条, 平均每个引物扩增的多态带数为8.41条, 多态性条带比率(PPB)为86.3%, 材料间遗传相似系数范围在0.6116到0.9290间。这说明鸭茅具有较丰富的遗传多样性。根据研究结果进行了聚类分析和主成分分析, 可将50份鸭茅材料分为5大类, 来自于相同洲的鸭茅能聚在一类, 中国和美国的鸭茅品种(系)能分别聚在同一类, 呈现出一定的地域性分布规律。并对鸭茅种质资源的收集保存提出建议。     

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.     

中国西南区扁穗牛鞭草种质遗传多样性的SRAP分析

本研究采用SRAP标记对主要来自中国西南地区(四川,重庆,贵州和云南)的43份扁穗牛鞭草种质资源的遗传多样性进行了分析。试验筛选出了11对引物组合对43份供试材料进行扩增,共获得153条带,其中多态性条带140条,多态性条带比率为91.50%,平均每对引物扩增出条带13.91,多态性条带12.73。实验数据结果表明,43份扁穗牛鞭草材料间的遗传相似系数(GS)为0.565～0.992,平均值为0.723,表现出了丰富的遗传多样性。聚类分析结果表明,各供试材料间的聚类与其地理来源以及形态特征类型具有一定的相关性。同时,主成分分析结果能够直观的反映了各种质间的遗传关系。5个扁穗牛鞭草地理类群间的分子方差分析(AMOVA)揭示了供试的扁穗牛鞭草总遗传变异的85.99%存在于类群内,仅有14.01%的变异存在于类群之间,类群间的分化系数ΦST=0.140。本研究结果为扁穗牛鞭草种质的收集、利用及育种提供了理论依据。     

基于SSR标记分析大豆疫霉根腐病抗源的遗传多样性

丰富的遗传多样性可为大豆育种提供宽阔的遗传基础,本研究基于35对SSR标记,对60份东北地区大豆疫霉根腐病抗性品种进行了遗传多样性分析,共检测到189个等位基因,平均每个位点等位变异数5.4个,多态性信息含量指数(PIC)为0.1550~0.8195,平均为0.6636;遗传相似系数的变异范围为0.31~0.74。利用5对高多态性SSR引物构建了60份抗性材料的指纹图谱,这5对SSR引物构建的指纹图谱可以将60份疫霉根腐病抗性材料逐一区分开。采用NTSYS2.10基于遗传距离的聚类分析,将60份抗性材料分为7个类群,其中78.33%的抗性品种(系)的遗传相似系数在0.45~0.74间,表明遗传差异相对较窄,品种间遗传多样性水平较低。聚类分析与群体遗传结构分析结果有部分重合,均反映出不同地区的抗性材料间存在一定的渗透和交流。     

黔南60份茶树种质资源遗传多样性的SSR分析

为探索黔南野生茶树种质资源的遗传多样性,利用SSR分子标记技术,对黔南60份茶树资源进行了DNA遗传多样性分析。结果表明:15对引物均显示多态性,基因多态性百分率为98.64%。15对SSR引物共扩增出147个观测等位基因和73.778 6个有效等位基因,平均每个引物扩增9.8个观测等位基因,4.918 6个有效等位基因。15个通用位点共产生280种基因型,平均每个位点18.7种基因型。遗传多态信息量变异范围为0.123 9~0.926 8,平均0.572 5,平均观测杂合度、平均期望杂合度和平均Shannons信息指数分别为0.470 0、0.602 3、1.464 4。经聚类分析后,60份材料间遗传相似系数在0.205 1~0.863 6之间,以平均遗传相似系数0.477 5为阈值,可将60份种质资源聚为8个类群,其在分子遗传水平上的分类结果与其材料来源分类的结果并不完全一致,而且材料来源地间遗传距离与地理距离不存在显著的相关性,有少部分同一来源的材料分散在各个类群中。研究认为,黔南茶树资源间的遗传差异较大,遗传基础较宽,具有丰富的遗传多样性。     

SRAP标记在落花生属种质资源遗传多样性上的利用

利用15对SRAP标记引物分析落花生属17份材料间的遗传变异情况。结果表明17份材料共检测出119个位点,其中多态性位点为100个,多态性百分率为89.00%,平均每对引物检测出6.67个多态性位点,扩增范围为5~10。材料间的遗传相似性范围为0.310~0.967,平均为0.666。基于遗传相似,利用UPGMA聚类分析表明,17份落花生可聚为3个类群,主成分分析析结果与聚类分析结果一致。研究结果证实了落花生种质资源遗传多样性丰富,为落花生育种工作奠定了基础。     

野生鸭茅种质遗传多样性的AFLP分析

本文应用扩增片段长度多态性（AFLP）技术,从64个引物对中筛选出9个,分别对37份鸭茅种质进行扩增,共得到400条清晰条带。其中多态性带336条,平均多态位点水平为84.0%,表现出较高的多态性。利用NTSYS-pc软件计算种质间的遗传距离,变化范围为0.0692～0.4214。用UPGMA方法进行聚类分析,在遗传相似系数为0.81水平上37份鸭茅种质可分为8个类群。用EIGEN方法进行主坐标分析,建立种质间相互关系的二维图。各种质在二维图中的分布与UPGMA聚类基本吻合。从分子水平分析,鸭茅遗传变异与染色体倍性和地理分布密切相关。     

扁蓿豆遗传多样性的等位酶研究

利用垂直聚丙烯酰胺凝胶电泳技术(SDS-PAGE)对24份扁蓿豆种质材料进行分析.结果表明:扁蓿豆具有较高的遗传多样性;4种等位酶共确定了6个等位酶位点,多态性百分率76%,平均每位点的等位基因的有效数目为1.492,平均预期杂合值为0.321;居群总遗传多样度为0.321,居群内的遗传多样性为0.273,居群内的遗传变异为81.9%,基因流(Nm)为1.4,居群间有一定的基因流动.UPGMA聚类分析结果表明:24个扁蓿豆天然居群可分为5大类,地理条件相似的居群优先聚为一类.     

Study of genetic diversity and relationships among accessions of foxtail millet [Setaria italica (L.) P. Beauv.] in Korea, China, and Pakistan using SSR markers

In this study, 28 simple sequence repeat (SSR) primer sets were used to analyze the genetic diversity, population structure, and genetic relationships among 37 accessions of foxtail millet from Korea, China and Pakistan. A total of 298 alleles were detected with an average allele number of 10.6 per locus among 37 foxtail millet accessions. The number of alleles per locus ranged from 2 (b226) to 20 (b236). Of the 298 alleles, 138 alleles (46.3%) were rare (frequency < 0.05), 152 alleles (51.0%) were detected at an intermediate frequency (range, 0.05?C0.50), and eight alleles (2.7%) were abundant (frequency > 0.50), respectively. The average gene diversity values were 0.652, 0.692, and 0.491 and polymorphic information content values were 0.621, 0.653, and 0.438, for accessions from Korea, China, and Pakistan, respectively. The accessions from China showed higher SSR diversity than those from Korea and Pakistan. A phylogenetic tree constructed using the un-weighted pair group methods with arithmetic mean algorithm revealed three major groups of accessions that were not congruent with geographical distribution patterns with a few exceptions. The lack of correlation between the accession clusters and their geographic location indicates that the diffusion of foxtail millet from China to Korea might have occurred through multiple routes. Our results provide support for the origin and diffusion route of foxtail millet in East Asia. This SSR-based assessment of genetic diversity, genetic relationships, and population structure among genetic resources of foxtail millet landraces will be valuable to foxtail millet breeding and genetic conservation programs in Korea.     

新疆甜高粱种质资源遗传多样性的SSR分析

选用50对SSR引物对新疆现有72份甜高粱种质资源进行遗传多样性分析。结果表明:有20对引物在72份甜高粱种质资源中表现为多态性。共检测到91个等位基因,每对引物可检测到的等位基因数目为2～5个,平均为3.45个。多态性信息量(PIC)的变动范围为0.2859～0.6652,平均为0.5057。72份甜高粱种质间的遗传相似系数变化范围为0.2001～1.000,平均值为0.5599。UPGMA聚类分析将72份材料划分为A、B两大类群,A群包括69份材料,而A群又被分成从Ⅰ到Ⅺ共11个亚群,B群包括3份材料,农艺性状近似的大多被聚到同一类群。     

Assessment of genetic diversity in broomcorn millet （Panicum miliaceum L.） using SSR markers

The genetic diversity of 118 accessions of broomcom millet （Panicum miliaceum L.）, collected from various ecological areas, was analyzed. Using 46 SSR （Simple Sequence Repeat） polymorphic markers from rice, wheat, oat and barley, a total of 226 alleles were found, which exhibited moderate level of diversity. The number of alleles per primer ranged from two to nine, with an average of 4.91. The range of polymorphism information content （PIC） was 0.2844）.980 （average, 0.793）. The expected heterozygosity （He） varied from 0.346 to 0.989, with an average of 0.834. The average coefficient of the genetic similarity of SSR markers among the 118 accessions was 0.609, and it ranged from 0.461 to 0.851. The UPGMA （Unweight Pair Group Method with Arithmetic Mean） clustering analysis at the genetic similarity value of 0.609 grouped the 118 accessions into five groups. Mantel test meant that geographical origin and genetic distance presented positive correlation. The clustering results were consistent with known information on ecological growing areas. The genetic similarity coefficient of the accessions in the Loess Plateau ecotype was significantly lower than those in the other ecotypes. It indicates that the highest level of genetic diversity occurred in the Loess Plateau, which is probably the original site of Panicum miliaceum.     

SSR标记在毛木耳种质资源遗传多样性研究中的应用

本研究利用基于毛木耳全基因组开发的SSR标记对27份毛木耳菌株（野生14株、栽培13株）的遗传多样性进行分析。首先随机选取3个菌株（2个野生菌株、1个栽培菌株）的DNA为模板,从144对SSR引物中筛选出扩增条带清晰、稳定性强、多态性丰富的引物24对。24对SSR引物共检测到116个多态性SSR片段,每对引物的多态性片段有3-7个,引物平均检测效率为4.83个,Shannon’s遗传多样性指数范围是0.866-1.885,多态性位点比率100%。供试菌株遗传相似系数范围是0.618-0.971,说明毛木耳种质资源具有丰富的遗传多样性。野生菌株与栽培菌株间平均遗传相似系数分别为0.746、0.779,说明毛木耳野生菌株遗传多样性更为丰富。经聚类分析,在遗传相似系数为0.680时,可将供试菌株分为无色（白色）类群Ⅰ和有色（浅黄色到红褐色）类群Ⅱ。遗传相似系数为0.704时,可将供试菌株中栽培菌株和野生菌株明显区分（14株野生菌株均在类群Ⅱ-2中,13株栽培菌株分别在类群Ⅰ和Ⅱ-1中）。本研究表明基于全基因组的SSR标记能从分子水平上揭示各菌株间的遗传差异,丰富毛木耳遗传多样性的研究手段,并为进一步进行毛木耳的品种选育、遗传学研究等提供有力手段。     

Genetic diversity analysis and transferability of cereal EST-SSR markers to orchardgrass (Dactylis glomerata L.)

The diversity and genetic relationships among 74 orchardgrass accessions were analyzed using cereal EST-SSRs and orchardgrass SSR markers in order to estimate genetic variability and compare the level of diversity. In total, 190 polymorphic bands were detected with an average of 6.3 alleles per SSR loci. The average polymorphic rate (P) for the species was 84.63%, suggesting a high degree of genetic diversity. The molecular variance analysis (AMOVA) showed that the proportion of variance explained by within- and among-geographical groups diversity was74.87% and 25.13%, respectively. The distinct geographical divergence of orchardgrass was revealed between Americas and Oceania. The ecogeographical conditions such as climate and soil, genetic drift and mating system could be the crucial factors for genetic divergence. Furthermore, the study also indicated that northern Africa, Europe and temperate Asia might be the diversity differentiation center of orchardgrass. The result will facilitate the breeding program and germplasm collection and conservation.     

High SSR diversity but little differentiation between accessions of Nordic timothy (Phleum pratense L.)

A large collection of genebank accessions of the hexaploid outcrossing forage grass species timothy (Phleum pratense L.) was for the first time analysed for SSR diversity on individual, population and regional level. Timothy is the most important forage grass species in the Nordic countries. Eighty-eight timothy accessions from Nordic countries and eight accessions around Europe were analysed with recently developed simple sequence repeat (SSR) markers. Timothy proved to be very polymorphic: the 13 selected SSRs amplified a total of 499 polymorphic alleles, the number of alleles per SSR locus varying from 15 to 74. Taking all SSR alleles together, the observed number in each accession ranged from 95 to 203. Levels of diversity were found to be significantly different between countries, vegetation zones and different cultivar types. However, the differentiation between accessions was low: most of the variation (94%) in the studied timothy material was due to variation within accessions and only 5% was between accessions and 1% between countries. Lack of geographical differentiation may reflect the outcrossing and hexaploid nature of timothy. Our results showed that neutral SSR markers are suitable for demonstrating levels of diversity but not alone adequate to resolve population structure in timothy. Nordic timothy material seems to be diverse enough for breeding purposes and no decline in the level of diversity was observed in varieties compared to wild timothy populations. Challenges in analysing SSR marker data in a hexaploid outcrosser were discussed.     

Expanding the repertoire of microsatellite markers for polymorphism studies in Indian accessions of mung bean (Vigna radiata L. Wilczek)

Limited availability of validated, polymorphic microsatellite markers in mung bean (Vigna radiata), an important food legume of India, has been a major hurdle towards its improvement and higher yield. The present study was undertaken in order to develop a new set of microsatellite markers and utilize them for the analysis of genetic diversity within mung bean accessions from India. A GA/CT enriched library was constructed from V. radiata which resulted in 1,250 putative recombinant clones of which 850 were sequenced. SSR motifs were identified and their flanking sequences were utilized to design 328 SSR primer pairs. Of these, 48 SSR markers were employed for assessing genetic diversity among 76 mung bean accessions from various geographical locations in India. Two hundred and thirty four alleles with an average of 4.85 alleles per locus were detected at 48 loci. The polymorphic information content (PIC) per locus varied from 0.1 to 0.88 (average: 0.49 per locus). The observed and expected heterozygosities ranged from 0.40 to 0.95 and 0.40 to 0.81 respectively. Based on Jaccard’s similarity matrix, a dendrogram was constructed using the unweighted pair-group method with arithmetic averages (UPGMA) analysis which revealed that one accession from Bundi, Rajasthan was clustered out separately while remaining accessions were grouped into two major clusters. The markers generated in this study will help in expanding the repertoire of the available SSR markers thereby facilitating analysis of genetic diversity, molecular mapping and ultimately broadening the scope for genetic improvement of this legume.     

A worldwide bread wheat core collection arrayed in a 384-well plate

Bread wheat (Triticum aestivum), one of the world’s major crops, is genetically very diverse. In order to select a representative sample of the worldwide wheat diversity, 3,942 accessions originating from 73 countries were analysed with a set of 38 genomic simple sequence repeat (SSR) markers. The number of alleles at each locus ranged from 7 to 45 with an average of 23.9 alleles per locus. The 908 alleles detected were used together with passport data to select increasingly large sub-samples that maximised both the number of observed alleles at SSR loci and the number of geographical origins. A final core of 372 accessions (372CC) was selected with this M strategy. All the different geographical areas and more than 98% of the allelic diversity at the 38 polymorphic loci were represented in this core. The method used to build the core was validated, by using a second set of independent markers [44 expressed sequence tag (EST)-SSR markers] on a larger sample of 744 accessions: 96.74% of the alleles observed at these loci had already been captured in the 372CC. So maximizing the diversity with a first set of markers also maximised the diversity at a second independent set of locus. To relate the genetic structure of wheat germplasm to its geographical origins, the two sets of markers were used to compute a dissimilarity matrix between geographical groups. Current worldwide wheat diversity is clearly divided according to wheat’s European and Asian origins, whereas the diversity within each geographical group might be the result of the combined effects of adaptation of an initial germplasm to different environmental conditions and specific breeding practices. Seeds from each accession of the 372CC were multiplied and are now available to the scientific community. The genomic DNA of the 372CC, which can be entirely contained in a 384-deep-well storage plate, will be a useful tool for future studies of wheat genetic diversity. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Isolation of EST-derived microsatellite markers for genotyping the A and B genomes of wheat

Genetic variation present in 64 durum wheat accessions was investigated by using three sources of microsatellite (SSR) markers: EST-derived SSRs (EST-SSRs) and two sources of SSRs isolated from total genomic DNA. Out of 245 SSR primer pairs screened, 22 EST-SSRs and 20 genomic-derived SSRs were polymorphic and used for genotyping. The EST-SSR primers produced high quality markers, but had the lowest level of polymorphism (25%) compared to the other two sources of genomic SSR markers (53%). The 42 SSR markers detected 189 polymorphic alleles with an average number of 4.5 alleles per locus. The coefficient of similarity ranged from 0.28 to 0.70 and the estimates of similarity varied when different sources of SSR markers were used to genotype the accessions. This study showed that EST-derived SSR markers developed in bread wheat are polymorphic in durum wheat when assaying loci of the A and B genomes. A minumum of ten EST-SSRs generated a very low probability of identity (0.36×10⁻¹²) indicating that these SSRs have a very high discriminatory power. EST-SSR markers directly sample variation in transcribed regions of the genome, which may enhance their value in marker-assisted selection, comparative genetic analysis and for exploiting wheat genetic resources by providing a more-direct estimate of functional diversity. Received: 19 December 2000 / Accepted: 17 April 2001