野生花生种质的SSR遗传多样性

以花生属(Arachis)6个区组24种(包括栽培种)84份种质为材料,用SSR技术对其亲缘关系和遗传多样性进行了分析.从206对SSR引物中筛选到59对能扩增出稳定的多态性条带的引物,这些引物能在花生属基因组DNA中扩增出1～6个DNA片段.结果表明,84份种质的遗传距离为0.04～0.93,平均为0.64,其中匍匐区组的A.appressipila的2份种质(G4与G5)的遗传距离最小(0.04),匍匐区组的A.rigonii(G14)与根茎区组的A.glabrata(G28)的遗传距离最大(0.93).聚类分析结果与花生属的区组分类基本一致,栽培种花生被聚在花生区组中,而且7份栽培种被聚在同一亚亚组中,相同植物学类犁(相当于变种)的材料均被分别聚在一起.异形花区组与直立区组的亲缘关系最近,与花生区组的亲缘关系较近的是匍匐区组.花牛区组的二倍体野生种A.villosa、A.duranensis和A.benensis与栽培种化生关系较近,可以作为桥梁物种来转移其他野生花生的优良基因.     

405份CIMMYT引进小麦种质的遗传多样性分析

为了明确国际玉米改良中心(CIMMYT)引进普通小麦种质材料的遗传多样性特点,为其利用提供参考依据,本研究从均匀分布于小麦基因组的420对SSR引物中选择出条带清晰、多态性较好的62对引物对引自CIMMYT的405份普通小麦种质系进行遗传多样性检测。结果表明,62对SSR引物在405份CIMMYT材料中共检测到198个等位变异,每对引物检测到等位变异的数目为2~8个,平均每对SSR引物能够检测到3.19个等位变异。单个SSR引物的PIC值介于0.03~0.79之间,平均值0.48。405份CIMMYT材料A、B、D基因组之间多态性位点数和等位变异数相差不大,PIC平均值B基因组(0.53)A基因组(0.52)D基因组(0.39)。聚类分析结果显示,62对SSR引物能够将405份CIMMYT材料区分开来,在0.1285遗传距离处将供试材料分为24个类群,类型较为丰富,不同类群的材料在农艺性状和品质性状上存在差异。     

棉花种间杂交渐渗系SSR及农艺性状分析

利用87对多态性SSR和EST-SSR引物,对57份棉属种间杂交渐渗系及其6个代表性血缘亲本进行了鉴定分析.共检测到540个等位变异,多态性等位变异占98.5%,EST-SSR变异占63.9%,41个SSR标记定位在棉花基因组的22条染色体上.种质间成对相似系数为0.553～0.937,平均为0.748.渐渗系的SSR聚类与种质材料的系谱来源基本吻合,而和农艺性状聚类结果相差很大,前者更能反映其亲缘关系.     

基于SSR标记分析小豆及其近缘植物的遗传关系

本研究利用87对SSR引物分析了80份栽培小豆(Vigna angularis)、22份野生小豆(V.angularis var.nipponensis)以及10份豇豆属(共7个种)近缘植物,旨在比较豇豆属不同种的遗传多样性,并分析种间的遗传关系.结果显示87对SSR引物在112份小豆及其近缘植物资源中共检测到667个等位变异.其中有75个、71个和82个SSR位点分别在栽培小豆、野生小豆和近缘植物中表现为多态.随机抽样分析发现,平均每SSR位点检测到的等位变异数目为近缘植物>野生小豆>栽培小豆,与多态信息含量(PIC)值一致,说明近缘植物及野生小豆中蕴含着丰富的遗传变异,是栽培小豆育种的重要基因来源.聚类分析显示,栽培小豆、野生小豆和近缘植物间的遗传分化比较明显,分别聚为三大类,其中栽培小豆的遗传背景与其生态环境相对应;近缘植物又可以分为三个亚类,亚类间的遗传距离与其亲缘关系相对应.本研究结果也说明利用SSR标记辅助豇豆属的种间分类是可行的.     

利用SSR标记分析云南、西藏和新疆小麦的遗传多样性

用185对SSR引物对52份中国西部特有小麦的遗传多样性进行了研究分析。在31份云南小麦材料中,共检测到488个等位变异,每一个SSR引物可检测到1至9个等位变异,平均为2.64个;平均PIC值为0.2764。在15份西藏小麦材料中,共检测到472个等位变异,每个引物可扩增出1到8个等位变异,平均为2.55个;平均PIC值为0.3082。在6份新疆小麦材料中,共检测到308个等位变异,每一个SSR引物可检测1到5个等位变异,平均为1.66个;平均PIC值为0.1944。185对SSR引物在云南、西藏和新疆小麦的21条染色体、7个部分同源群和3个染色体组上检测到的等位位点的多态性存在明显差异。云南、西藏和新疆小麦均以3B染色体较高,而1D染色体最低;在7个部分同源群中,均以第三部分同源群最高,第六部分同源群最低;在A、B和D染色体组上,均以B染色体组最高,D染色体组最低,A染色体组居中。利用185对SSR引物计算了云南、西藏和新疆小麦群体内及其群体间的遗传距离(GD)和平均遗传距离,结果显示,西藏小麦和云南小麦群体内的平均遗传距离要高于新疆小麦,而云南小麦和西藏小麦间的平均遗传距离低于两者与新疆小麦的平均遗传距离。聚类分析结果也表明,云南小麦和西藏小麦的亲缘关系较近,但两者与新疆小麦的亲缘关系相对较远。     

广东省猴耳环遗传多样性研究

为了解猴耳环(Archidendron clypearia)种质资源的遗传多样性,以广东省12个野生猴耳环群体的146份种质资源为材料,采用SSR分子标记技术对其遗传多样性和亲缘关系进行分析。结果表明,21对SSR引物共检测到249个等位基因,平均每对SSR引物检测的等位基因数(Na)为11.857,有效等位基因数(Ne)为3.500,期望杂合度(He)为0.718,多态信息含量(PIC)为0.676;12个群体中博罗群体的Shannon多样性指数(I=0.528)和有效等位基因数(Ne=0.716)均最大,是遗传多样性最丰富的群体;群体间的遗传分化系数为0.071,AMOVA分析表明,猴耳环的遗传变异主要在群体内(97%),群体内的遗传分化大于群体间。聚类分析表明,遗传系数在0.16时,可将12个群体分为6大类,与主坐标分析的结果大致相同。这为发掘、利用与保护猴耳环群体种质资源,开展猴耳环优良品种的遗传育种提供重要的理论依据。     

白及种质资源及其近缘种的SSR指纹图谱研究

白及(Bletilla striata Rchb.)为中国珍稀濒危药用植物,野生资源枯竭,混伪种繁多。该研究基于SSR标记技术对16个地区48份白及(Bletilla striata Rchb.)样品和4份黄花白及(Bletilla ochracea Schltr.)、4份小白及(Bletilla formosana(Hayata)Schltr.)、4份华白及(Bletilla sinensis(Rolfe)Schltr.)样品进行遗传多样性分析,构建白及种质资源的SSR指纹图谱,并对60份白及样品进行种质资源鉴定,分析白及属种内及其种间的遗传分化特征。结果表明:(1)20对白及SSR引物均能在4个白及属植物中成功扩增,条带清晰且多态性丰富,每对引物的复等位基因数(Na)在4~9之间,等位基因数(Na)总和为127,平均为6.35。(2)筛选出基因型丰富、多态性信息含量(PIC)高的7对白及SSR引物(BJSSR01、BJSSR14、BJSSR15、BJSSR16、BJSSR18、BJSSR19、BJSSR22)构建的白及SSR指纹图谱能将白及属各种质资源清楚分开。(3)白及属在种间水平均有较高的遗传多样性(Na=6.35,I=1.429 1,h=0.706 8),物种间遗传分化强烈(Gst=0.44),物种间的基因流较弱(Nm=0.475 3)。(4)UPGMA聚类分析表明,60份供试样品明显聚为4大支,同一物种的个体首先聚在一起,这与形态学分类基本一致;不同来源地的样品种内和种间的亲缘关系有明显差异,地理距离较近的白及样品具有较近的遗传关系。     

内蒙古3种生态型扁蓿豆遗传多样性与亲缘关系的分析

采用SSR分子标记结合21个表型性状对来自内蒙古3种生态型扁蓿豆种质资源的遗传多样性和亲缘关系进行了分析.结果表明:3种生态型的22份扁蓿豆材料在考察的4个质量性状上差异明显;在17个数量性状上,总体变异程度为黄花型扁蓿豆>扁蓿豆>细叶扁蓿豆;表型性状的遗传相似系数在31.59～113.27间,变异系数为43.30%.SSR分析结果显示,18对引物平均多态性比率为80.09%,引物平均等位位点数为6.06,位点多态性信息含量平均为0.32,遗传相似系数在0.37～0.47间,变异系数为61.80%.表型性状聚类、SSR分子标记聚类及主成分分析结果均显示,黄花型扁蓿豆和扁蓿豆的亲缘关系较近,与细叶扁蓿豆的亲缘关系较远.     

广西武宣濠江流域普通野生稻居群遗传多样性及保护研究

选用平均分布于水稻基因组的24对SSR引物,对沿河分布最长的广西武宣濠江流域的12个普通野生稻居群343份材料的遗传结构进行研究.结果表明:(1)该地普通野生稻遗传多样性丰富.24个位点共检测到206个等位变异,平均等位变异数A＝8.7083,有效等位变异数Ae＝3.7117;(2)该地普通野生稻居群具有较高的遗传分化和一定频率的基因流.群体遗传分化系数Gst＝0.2659,基因流Nm＝0.6901,表明26.59%的遗传变异存在于居群间;(3)SSR标记使普通野生稻居群中一些稀有等位变异得以显现.206个等位变异中,65个等位变异仅出现在1个或2个居群中,且频率较低,其中12个等位变异只出现在居群B中;(4)通过聚类分析和主坐标分析(PCO),下游居群A和B遗传关系较近,中游居群C比较独特,单独成为一类,中游居群D、E、F和G遗传关系较近,中游居群H、I和J及上游居群K和L遗传关系较近.根据上述分析结果,建议对濠江下游和中游具有代表性的居群(即居群B、D和H)的普通野生稻进行重点保护.     

设施用厚皮甜瓜品种SSR标记遗传多样性分析

使用分布于甜瓜12条染色体上的72对SSR引物,对我国中东部设施内栽培的30个厚皮甜瓜品种进行分析;56对SSR引物在30个品种间表现为多态性。共检测到138个等位变异,每对引物的等位变异数变幅为2～6个,平均为2.6个。有效等位变异为86.16个,平均为2.25。每个SSR位点的多态性信息量(PIC)变化范围为0.045～0.725,平均为0.390。30个品种间遗传相似系数变幅为0.274～0.974之间,平均值为0.665,且90.4%的供试品种其遗传相似系数在0.474～0.824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,我国中东部设施适宜种植的甜瓜品种遗传多样性不够丰富,多数品种间的亲缘关系较近,欲进一步提高中东部地区设施甜瓜产量和品质还需要拓宽亲本选择范围,扩大遗传背景。     

RAPD and ISSR fingerprints as useful genetic markers for analysis of genetic diversity, varietal identification, and phylogenetic relationships in peanut (Arachis hypogaea) cultivars and wild species.

Abstract: Twenty-one random and 29 SSR primers were used to assess genetic variation and interrelationships among subspecies and botanical varieties of cultivated peanut, Arachis hypogaea (2n = 4x = 40), and phylogenetic relationships among cultivated peanut and wild species of the genus Arachis. In contrast with the previous generalization that peanut accessions lack genetic variation, both random and SSR primers revealed 42.7 and 54.4% polymorphism, respectively, among 220 and 124 genetic loci amplified from 13 accessions. Moreover, the dendrograms based on RAPD, ISSR, and RAPD + ISSR data precisely organized the five botanical varieties of the two subspecies into five clusters. One SSR primer was identified that could distinguish all the accessions analysed within a variety. Although the polymorphic index content varied from 0.1 to 0.5 for both ISSR and RAPD markers, primer index values were substantially higher for RAPD primers (0.35-4.65) than for SSR primers (0.35-1.73). It was possible to identify accessions, particularly those of divergent origins, by RAPD and (or) ISSR fingerprints. Based on these results, marker-based genetic improvement in A. hypogaea appears possible. None of the 486 RAPD and 330 ISSR amplification products were found to be commonly shared among 13 species of section Arachis and one species each of sections Heteranthae, Rhizomatosae, and Procumbentes. Dendrograms constructed from RAPD, ISSR, and RAPD + ISSR data showed overall similar topologies. They could be resolved into four groups corresponding to the species grouped in four taxonomic sections. The present results strongly support the view that Arachis monticola (2n = 4x = 40) and A. hypogaea are very closely related, and indicate that A. villosa and A. ipaensis are the diploid wild progenitors of these tetraploid species.     

Seed storage protein variation in Arachis species.

Seventy-two accessions, representing 22 species from sections Arachis, Erectoides, Extranervosae, and Triseminalae of the genus Arachis, were screened for seed storage protein polymorphism. Variation was detected between sections, between genome types, between species, and in some cases between different accessions of the same species or different seeds of the same accession. Arachis duranensis and one accession of A. cardenasii were found to have identical protein patterns. The greatest dissimilarity was found between species of the section Extranervosae and species of the section Triseminalae. Those of section Erectoides showed much similarity with some species of section Arachis. Protein polymorphism was shown to distinguish the two subspecies of A. hypogaea (fastigiata and hypogaea) in 27 of 28 cases. The seed protein profile of A. monticola was a combination of seed protein profiles from the two A. hypogaea subspecies. The relatedness between the various species was calculated and those that had the greatest similarity with A. hypogaea were A. spegazzinii and A. batizocoi.     

Taxonomic relationships among Arachis sect. Arachis species as revealed by AFLP markers.

Cultivated peanut, Arachis hypogaea L., is a tetraploid (2n = 4x = 40) species thought to be of allopolyploid origin. Its closest relatives are the diploid (2n = 2x = 20) annual and perennial species included with it in Arachis sect. Arachis. Species in section Arachis represent an important source of novel alleles for improvement of cultivated peanut. A better understanding of the level of speciation and taxonomic relationships between taxa within section Arachis is a prerequisite to the effective use of this secondary gene pool in peanut breeding programs. The AFLP technique was used to determine intra- and interspecific relationships among and within 108 accessions of 26 species of this section. A total of 1328 fragments were generated with 8 primer combinations. From those, 239 bands ranging in size from 65 to 760 bp were scored as binary data. Genetic distances among accessions ranged from 0 to 0.50. Average distances among diploid species (0.30) were much higher than that detected between tetraploid species (0.05). Cluster analysis using different methods and principal component analysis were performed. The resulting grouping of accessions and species supports previous taxonomic classifications and genome designations. Based on genetic distances and cluster analysis, A-genome accessions KG 30029 (Arachis helodes) and KSSc 36009 (Arachis simpsonii) and B-genome accession KGBSPSc 30076 (A. ipaensis) were the most closely related to both Arachis hypogaea and Arachis monticola. This finding suggests their involvement in the evolution of the tetraploid peanut species.     

用SSR和AFLP技术分析花生抗青枯病种质遗传多样性的比较

由Ralstonia solanacearum E.F.Smith引起的青枯病是若干亚洲和非洲国家花生生产的重要限制因子,利用抗病品种是防治这一病害最好的措施。虽然一大批抗青枯病花生种质资源材料已被鉴定出来,但对其遗传多样性没有足够的研究,限制了在育种中的有效利用。本研究以31份对青枯病具有不同抗性的栽培种花生种质为材料,通过简单序列重复(SSR)和扩增片段长度多态性(AFLP)技术分析了它们的遗传多样性。通过78对SSR引物和126对AFLP引物的鉴定,筛选出能显示抗青枯病种质多态性的SSR引物29对和AFLP引物32对。所选用的29对多态性SSR引物共扩增91条多态性带,平均每对引物扩增3.14条多态性带;32对多态性AFLP引物共扩增72条多态性带,平均扩增2.25条多态性带。在所筛选引物中,4对SSR引物(14H06,7G02,3A8,16C6)和1对AFLP引物(P1M62)检测花生多态性的效果优于其他引物。SSR分析获得的31个花生种质的遗传距离为0.12-0.94,平均为0.53,而AFLP分析获得的遗传距离为0.06～0.57,平均为0.25,基于SSR分析的遗传距离大于基于AFLP分析的遗传距离,疏枝亚种组的遗传分化相对大于密枝亚种组。基于两种分析方法所获得的聚类结果基本一致,但SSR数据聚类结果与栽培种花生的形态分类系统更为吻合。根据分析结果,对构建青枯病抗性遗传图谱群体的核心亲本和抗性育种策略提出了建议。     

Use of single-primer DNA amplifications in genetic studies of peanut (Arachis hypogaea L.)

A recent approach to detecting genetic polymorphism involves the amplification of genomic DNA using single primers of arbitrary sequence. When separated electrophoretically in agarose gels, the amplification products give banding patterns that can be scored for genetic variation. The objective of this research was to apply these techniques to cultivated peanut (Arachis hypogaea L.) and related wild species to determine whether such an approach would be feasible for the construction of a genetic linkage map in peanut or for systematic studies of the genus. Two peanut cultivars, 25 unadapted germplasm lines of A. hypogaea, the wild allotetraploid progenitor of cultivated peanut (A. monticola), A. glabrata (a tetraploid species from section Rhizomatosae), and 29 diploid wild species of Arachis were evaluated for variability using primers of arbitrary sequence to amplify segments of genomic DNA. No variation in banding pattern was observed among the cultivars and germplasm lines of A. hypogaea, whereas the wild Arachis species were uniquely identified with most primers tested. Bands were scored (+/–) in the wild species and the PAUP computer program for phylogenetic analysis and the HyperRFLP program for genetic distance analysis were used to generate dendrograms showing genetic relationships among the diploid Arachis species evaluated. The two analyses produced nearly identical dendrograms of species relationships. In addition, approximately 100 F₂ progeny from each of two interspecific crosses were evaluated for segregation of banding patterns. Although normal segregation was observed among the F₂ progeny from both crosses, banding patterns were quite complex and undesirable for use in genetic mapping. The dominant behavior of the markers prevented the differentiation of heterozygotes from homozygotes with certainty, limiting the usefulness of arbitrary primer amplification products as markers in the construction of a genetic linkage map in peanut.     

Abundant Microsatellite Diversity and Oil Content in Wild Arachis Species

The peanut (Arachis hypogaea) is an important oil crop. Breeding for high oil content is becoming increasingly important. Wild Arachis species have been reported to harbor genes for many valuable traits that may enable the improvement of cultivated Arachis hypogaea, such as resistance to pests and disease. However, only limited information is available on variation in oil content. In the present study, a collection of 72 wild Arachis accessions representing 19 species and 3 cultivated peanut accessions were genotyped using 136 genome-wide SSR markers and phenotyped for oil content over three growing seasons. The wild Arachis accessions showed abundant diversity across the 19 species. A. duranensis exhibited the highest diversity, with a Shannon-Weaver diversity index of 0.35. A total of 129 unique alleles were detected in the species studied. A. rigonii exhibited the largest number of unique alleles (75), indicating that this species is highly differentiated. AMOVA and genetic distance analyses confirmed the genetic differentiation between the wild Arachis species. The majority of SSR alleles were detected exclusively in the wild species and not in A. hypogaea, indicating that directional selection or the hitchhiking effect has played an important role in the domestication of the cultivated peanut. The 75 accessions were grouped into three clusters based on population structure and phylogenic analysis, consistent with their taxonomic sections, species and genome types. A. villosa and A. batizocoi were grouped with A. hypogaea, suggesting the close relationship between these two diploid wild species and the cultivated peanut. Considerable phenotypic variation in oil content was observed among different sections and species. Nine alleles were identified as associated with oil content based on association analysis, of these, three alleles were associated with higher oil content but were absent in the cultivated peanut. The results demonstrated that there is great potential to increase the oil content in A. hypogaea by using the wild Arachis germplasm.     

江西野生大豆遗传多样性分析

利用48个SSR分子标记分析了192份采集于江西49个县（区）的野生大豆遗传多样性。结果表明,共扩增出等位基因343个,平均每对引物扩增出7.2个等位基因,平均基因遗传多样性指数0.7369,平均多态性信息含量（PIC）0.7060,表明江西野生大豆具有较为丰富的遗传多样性。不同纬度和不同海拔的野生大豆其遗传多样性不同,高纬度及低海拔地区野生大豆遗传多样性要高。192份野生大豆可聚类成三大类,聚类结果与地理来源较为一致。     

Start codon targeted polymorphism for evaluation of functional genetic variation and relationships in cultivated peanut (<Emphasis Type="Italic">Arachis</Emphasis><Emphasis Type="Italic">hypogaea</Emphasis> L.) genotypes

Cultivated peanut possesses an extremely narrow genetic basis. Polymorphism is considerably difficult to identify with the use of conventional biochemical and molecular tools. For the purpose of obtaining considerable DNA polymorphisms and fingerprinting cultivated peanut genotypes in a convenient manner, start codon targeted polymorphism technique was used to study genetic diversity and relatedness among 20 accessions of four major botanical varieties of peanut. Of 36 primers screened, 18 primers could produce unambiguous and reproducible bands. All 18 primers generated a total of 157 fragments, with a mean of 8.72 ranging from 4 to 17 per primer. Of 157 bands, 60 (38.22%) were polymorphic. One to seven polymorphic bands were amplified per primer, with 3.33 polymorphic bands on average. Polymorphism per primer ranged from 14.29 to 66.67%, with an average of 36.76%. The results revealed that not all accessions of the same variety were grouped together and high genetic similarity was detected among the tested genotypes based on cluster analysis and genetic distance analysis, respectively. Further, accession-specific markers were observed in several accessions. All these results demonstrated the following: (1) start codon targeted polymorphism technique can be utilized to identify DNA polymorphisms and fingerprint cultivars in domesticated peanut, and (2) it possesses considerable potential for studying genetic diversity and relationships among peanut accessions.     

Variation in chromosomal DNA associated with the evolution of Arachis species.

The 2C nuclear DNA amounts were determined for 99 accessions, representing 23 Arachis species from 8 of 9 taxonomic sections, and two synthetic amphidiploids. Mean 2C DNA amounts varied by 15.20%, ranging from 10.26 to 11.82 pg, between accessions of Arachis hypogaea (2n = 4x = 40). Nuclear DNA content variation (5.33-5.91 pg) was also detected among Arachis duranensis (2n = 2x = 20) accessions. The intraspecific variation in the two species may have resulted from indirect selection for favourable genome sizes in particular environmental conditions. The accessions belonging to A. hypogaea ssp. hypogaea (mean value 11.27 pg) with longer life cycle had significantly larger mean DNA content than the accessions of A. hypogaea ssp. fastigiata (mean value 10.97 pg). For 20 diploid (2n = 2x = 20) species of the genus, 2C nuclear DNA amounts ranged from approximately 3 to 7 pg. The diploid perennial species of section Arachis have about 12% more DNA than the annual species. Comparisons of DNA amounts show that evolutionary rating is not a reliable guide to DNA amounts in generic sections of the genus; lower DNA values with evolutionary advancement were found in sections Heteranthae and Triseminatae, but the same was not true for sections Arachis and Caulorrhizae. Similarly, there is evidence of significant differences in DNA content between 4 ancient sections (Procumbentes, Erectoides, Rhizomatosae, and Extranervosae) of the genus. The occurrence of genome size plasticity in both A. duranensis and A. hypogaea provides evidence that A. duranensis could be one of the diploid progenitors of A. hypogaea. The DNA content in the two synthetic amphidiploids corresponded to the sum value estimated for parental species. Key words : Arachis species, genome size, Arachis hypogaea, Arachis duranensis, intraspecific variation.