16份石榴RAPD扩增产物的两种电泳方法检测及其序列特征

本实验以16个石榴品种为实验材料,筛选出10个重复性及多态性均较好的引物进行RAPD分析。分别采用琼脂糖凝胶以及聚丙烯酰胺凝胶(PAGE)电泳检测方法对PCR扩增结果进行检测并对其结果进行比较,结果显示,两种电泳方式均能得到较为清晰的扩增条带,且两种电泳方式获得的条带总数及多态性条带数均有所不同,琼脂糖凝胶电泳方法共检测出76条带,其中有43条为多态性谱带,多态性比率为56.4%;而在PAGE电泳方法共检测出123条谱带,多态性谱带数为87条,多态性比率为70.95%。PAGE电泳方法检测出的条带数约为琼脂糖凝胶电泳方法检测出条带数的1.5倍。基于两种电泳方法所得RAPD标记的多态性位点,利用NYSYS软件计算遗传相似系数,并构建遗传关系聚类图,分析结果显示,石榴遗传多样性丰富,两种电泳方法所得聚类结果大致相同,可以利用RAPD分子标记及两种电泳检测方法对不同数量的石榴进行分子水平的品种鉴定和遗传多样性的分析。同时通过对来自几个引物随机挑选的17个片段进行克隆,测序结果显示17个片段都是对应引物的RAPD扩增产物,其中有3条是编码蛋白的基因片段,表明了RAPD不仅扩增基因组上的非编码蛋白序列,同时也可以扩增编...     

利用RAPD和AFLP标记分析烟草种质资源的遗传多样性

从200条10 bp的RAPD引物中筛选获得28条多态性引物，对48份烟草材料的基因组DNA进行扩增。共获得184条DNA扩增片断带，其中多态性带86条，平均多态检出率为46.7%。用4对AFLP选择性扩增引物对48份烟草材料进行选择性扩增，共获得321条扩增带，其中174条具有多态性，平均多态检出率为54.2%。根据RAPD和AFLP标记的结果，采用UPGMA法进行聚类分析，两种方法获得了相似但不完全相同的结果，两者都可以将48份烟草资源分为两大类群，即黄花烟草(Nicotiana rustica)群和普通烟草(N.tobacum)群，普通烟草可进一步分为4组；48份材料的遗传距离变幅在1.4～11.0之间，其聚类结果与理论上所期望的结果基本一致，AFLP标记技术比RAPD标记技术能更好地从分子水平上揭示烟草种质资源的遗传背景、亲缘关系及演化规律。     

RAPD和ISSR标记对甜瓜种质遗传多样性的研究

利用RAPD和ISSR两种分子标记技术对37份甜瓜(CucumismeloL.)种质进行了遗传多样性研究.在供试材料中筛选到具有多态性的RAPD引物21个,ISSR引物10个(其中ISSR-2与ISSR-4等量混合组成ISSR-10引物对).RAPD引物共扩增出多态性带106条,多态性条带比率(PPB)为58.62%,平均多态信息量(PIC)为0.47;ISSR引物共扩增出多态性带73条,PPB值为65.51%,平均PIC值为0.53.根据两种标记的结果,采用UPGMA聚类分析,将供试材料分为两大类群野生甜瓜和栽培甜瓜;栽培甜瓜又分为厚皮甜瓜和薄皮甜瓜两大亚群,各野生甜瓜种质之间的遗传距离较大,这与其分类地位基本一致.两种分子标记的分析结果呈极显著正相关(r=0.62>r=0.01).研究表明,RAPD和ISSR标记可用于甜瓜种质遗传多样性的研究.     

47份水稻品种资源的ISSR遗传多样性分析

为研究广东省惠州市种植的常规水稻品种的遗传多样性,本实验利用ISSR标记对47份水稻品种资源进行遗传多样性检测。从49条引物中筛选出5条重复性好,条带清晰的引物进行PCR扩增,共扩增出53条带,每个引物可以扩增出9～13条带,平均为10．6条,其中47条具有多态性,比率为88．7％。不同水稻品种间遗传相似系数变幅为0．319～0．936,平均达0．691,说明ISSR标记能够揭示材料间较高的遗传多样性。通过聚类,从分子水平对水稻品种资源的遗传关系进行分析,并对47份水稻品种资源进行分类,ISSR标记能将47份水稻品种完全区分开,为水稻品种资源的研究利用提供参考。     

野生斑鳠西江地理群的遗传多样性分析

为了解斑鳠(Mystus guttatus)不同地理种群之间的遗传分化程度,基于RAPD技术,分析了珠江水系西江段野生斑鳠的遗传多样性。利用20个随机引物对30个斑鳠个体的基因组DNA进行了PCR扩增,共扩增出3210条DNA片段,平均每个个体扩增出107条带。在检测到的107个位点中,多态位点数为48个,占44.9%,仅有一个引物S30没有扩增出多态带。个体间最大的遗传距离0.2804,最小的遗传距离0.0467,平均遗传距离0.1526±0.037,种群内个体间平均的相似率为84.7±3.7%。     

草莓品种的亲缘关系及其不同继代次数的叶片组培苗的RAPD分析

从100个随机引物中筛选出10个引物,对16个草莓品种进行RAPD分析。结果显示,10个引物共扩增出73条DNA谱带,其中多态条带49条,多态性程度为67.13%;遗传距离D值在0.40水平上能将供试材料聚为3类,表明RAPD能很好地鉴别草莓品种之间的遗传关系。利用前面筛选的10个随机引物和其他20个随机引物对不同继代次数的草莓组培继代苗进行了RAPD分析,其特征带型表现一致,未发现变异,说明草莓叶盘离体继代培养30次以内能稳定遗传。     

中国南瓜种质资源农艺性状与RAPD标记分析

本研究利用形态学标记和RAPD分子标记同时对70份来自我国不同地区的中国南瓜种质进行遗传多样性和亲缘关系分析。在所观察的56个有差异的农艺性状中,变异系数从6.60%~262.22%,平均变异系数为37.50%。从150个随机引物中筛选出21个进行RAPD分析,结果表明,在检测到的167条带中有130条具有多态性,多态带比率为73.23%;平均每个引物检测到的条带数多达8条,平均Shannon信息指数(Ⅰ)为0.268。基于农艺性状的聚类分析将70份中国南瓜种质分为七类。基于RAPD标记的聚类分析将70份中国南瓜种质分为五类,其聚类结果与形态特征有一定的相关性。但两种聚类结果均无法从地理来源上进行区分。     

泰国产方斑东风螺养殖群体遗传多样性的RAPD分析

用随机扩增多态性DNA(RAPD)技术对泰国产方斑东风螺养殖群体的遗传多样性进行检测,从100个随机引物中筛选出21个引物对方斑东风螺的DNA进行扩增,结果表明:21个引物共检测到222条清晰且重复性好的条带,每个引物可扩增出4~16条带,分子量在200~2200bp之间,其中多态位点为156个,占70.27%;群体的Shannon多样性指数为0.2818,Nei基因多样性指数为0.2491;个体间最大遗传距离为0.291,最小遗传距离为0.066。通过与其他贝类遗传多样性的研究结果比较,可初步判断泰国产方斑东风螺养殖群体的遗传多样性比较丰富。     

应用RAPD技术研究4种鲍的亲缘关系

应用RAPD技术研究 4种鲍的亲缘关系结果表明 ,4种鲍群体中 2 0个有效引物共扩增出 5 38条DNA带 ,平均每个引物扩增的条带数为 2 6 .9;扩增的多态性条带数 136 ,平均每个引物扩增的多态性条带数为 6 .8;多态位点百分数为 2 5 .3%。盘鲍群体和皱纹盘鲍群体之间遗传距离与遗传一致度分别为 0 .2 8和 0 .72 ,杂色鲍群体和九孔鲍群体之间遗传距离与遗传一致度分别为 0 .32和 0 .6 8。聚类分析把盘鲍群体和皱纹盘鲍群体聚为 1组 ,二者亲缘关系较近 ;杂色鲍群体和九孔鲍群体聚为 1组 ,二者亲缘关系也较近     

广西地方葛根种质资源遗传多样性的SCoT分析

为了解广西地方葛根种质资源之间的亲缘关系,本研究以SCoT分子标记对44份广西葛根种质资源的基因组DNA进行PCR扩增,并进行遗传多样性分析。结果表明,25条SCoT引物共扩增出223个条带,其中具有多态性的条带有194条,平均每条引物获得7.76个多态性条带,多态性比例为86.99%。聚类分析表明,44份葛根种质资源的遗传相似系数在0.587~0.982之间。在系数0.65处,44份葛根分为两大类,37号材料单独成为一类;在系数为0.74处,可聚为六类。综上所述,SCoT分子标记适用于葛根种质资源遗传多样性分析,本研究结果为广西葛根种质资源鉴定评价和新品种选育奠定了一定的理论基础。     

Measurement of genetic dissimilarity in fieldpea (<Emphasis Type="Italic">Pisum sativum</Emphasis> L.) genotypes using RAPD markers

The present investigation was carried out on fifteen germplasm lines of Pisum sativum L. were used for characterization using Randomly Amplified Polymorphic DNA (RAPD) markers. While 12 random primers were taken, out of them 11 primers gave amplification. These primers gave a total of 133 bands out of which 106 were polymorphic. Genetic similarities of the RAPD profiles were estimated by using Jaccard’s coefficient with NTSYSpc 2.0 software. The similarity index values ranged from 0.263 to 0.793 indicating the presence of enormous genetic diversity at molecular level. A dendrogram generated by cluster analysis divided fifteen fieldpea genotypes into two Groups A and B. Major Group A have five genotypes and major Group B have nine genotypes.     

Genetic Diversity Among Brazilian Cultivars and Landraces of Tomato Lycopersicon Esculentum Mill. Revealed by RAPD Markers

Random amplified polymorphic DNA markers (RAPD) were used to estimate the variability of 35 tomato accessions (Lycopersicon esculentum Mill.). A total of 257 reproducibly scorable bands were obtained from 20 primers, 78.6% of which were polymorphic. The percentage distribution of RAPD markers shows a bimodal distribution, and the frequency of rare alleles is similar in commercial and landrace accessions. Genetic distances among accessions were calculated and a dendrogram showing the genetic relationships among them was constructed allowing for the separation of four groups. Twenty out of 23 Brazilian landraces fell within one group, whereas commercial cultivars were distributed in the four groups. AMOVA analysis of RAPD data showed that, despite the high within Brazilian landraces and commercial cultivars variation, these two groups are significantly different, indicating that landraces can be a source of variation for breeding programs.     

Characterization of Some Italian Common Bean (<Emphasis Type="Italic">Phaseolus Vulgaris</Emphasis> L.) Landraces by RAPD,Semi-random and ISSR Molecular Markers

Randomly amplified polymorphic DNA (RAPD), inter-simple sequence repeat (ISSR) and a semi-random PCR system were used to analyze the genetic diversity of 16 Italian common bean landraces and their relationship to four commercial cultivars. Of the primers tested, 8 ISSR, 6 RAPD and 7 semi-random primers produced polymorphic and reproducible DNA fragments. A higher proportion of polymorphic bands were observed using ISSR (85%) and semi-random (90%) primers than RAPD (69%) method. The combination of any two semi-random markers allowed the identification of all 20 bean genotypes. In contrast ISSR (except for primer (CAC)₃GC) and RAPD markers appeared to be less informative as more than two markers were necessary to achieve the same diagnostic level. Moreover, 7 ISSR, 2 RAPD and 8 semi-random exclusive bands were identified as putative population-specific markers. Semi-random and ISSR derived dendrograms showed similar tendencies in terms of genetic relatedness, whereas clustering of genotypes within groups was not similar when compared with the RAPD technique. Despite the different ability to resolve genetic variation among the investigated landraces, two major clusters with less than 60% (ISSR) and 40% (RAPD and semi-random) genetic similarity were formed with all three marker systems. The two groups were correlated with the phaseolin patterns and seed size of the landraces. The analysis showed that the cultivar ȁ8Lingua di Fuocoȁ9 and most of the landraces (13 out of 16) collected in Italy belong to the Andean gene pool, whereas only the three populations from Pratomagno belong to the Middle American gene pool.     

RAPD and ISSR molecular markers in <Emphasis Type="Italic">Olea europaea</Emphasis> L.: Genetic variability and molecular cultivar identification

Thirty Portuguese and eight foreign olive (Olea europaea L.) cultivars were screened using Random Amplified Polymorphic DNA (RAPD) and Inter-Simple Sequence Repeat (ISSR) markers. Twenty RAPD primers amplified 301 reproducible bands of which 262 were polymorphic; and 17 ISSR primers amplified 204 bands of which 180 were polymorphic. The percentage of polymorphic bands detected by ISSR and RAPD was similar (88 and 87%, respectively). The genetic variability observed was similar in the Portuguese and foreign olive cultivars. Seven ISSR and 12 RAPD primers were able to distinguish individually all 38 olive cultivars. Twenty specific molecular markers are now available to be converted into Sequence Characterised Amplified Region (SCAR) markers. Relationships among Portuguese and foreign cultivars is discussed.     

The genetic characterization of Turkish watermelon (<Emphasis Type="Italic">Citrullus lanatus</Emphasis>) accessions using RAPD markers

Genetic diversity of the Turkish watermelon genetic resources was evaluated using different Citrullus species, wild relatives, foreign landraces, open pollinated (OP) and commercial hybrid cultivars by RAPD markers. The germplasm was consisted of 303 accessions collected from various geographical regions. Twenty-two of 35 RAPD primers generated a total of 241 reproducible bands, 146 (60.6%) of which were polymorphic. Based on the RAPD data the genetic similarity coefficients were calculated and the dendrogram was constructed using UPGMA (Unweighted pair-group method with arithmetic average). Cluster analysis of the 303 accessions employing RAPD data resulted in a multi-branched dendrogram indicating that most of the Turkish accessions belonging to var. lanatus of Citrullus lanatus (Thunb.) Matsum et Nakai were grouped together. Accessions of different Citrullus species and Praecitrullus fistulosus (Stocks) Pangalo formed distant clusters from C. lanatus var. lanatus. Among 303 accessions, a subset of 56 accessions was selected representing different groups and a second dendrogram was constructed. The genetic similarity coefficients (GS) within the Turkish accessions were ranged from 0.76 to 1.00 with 0.94 average indicating that they are closely related. Taken together, our results indicated that low genetic variability exist among the watermelon genetic resources collected from Turkey contrary to their remarkable phenotypic diversity.     

海南荔枝(Litchi chinensis Sonn.)主要栽培品种的RAPD分析

从80个随机引物中筛选出12条重复性好的多态性引物，对包括15个栽培品种在内的22份海南荔枝(Litchi chinensis Sonn.)资源进行RAPD分析，扩增的多态性片段比例为64.1%, 平均为31.1%; 遗传距离在0.0000~0.6180之间，平均0.3732。UPGMA聚类分析显示，22份资源主要分为2类，部分供试品种存在明显的同名异物现象。     

RAPD markers in the analysis of genetic diversity among common bean germplasm from Central Himalaya

A set of 99 common bean germplasm collected from central Himalaya was investigated for their genetic variability using random amplified polymorphic DNA (RAPD) markers. Ten oligonucleotide primers, selected from 60 initially screened, generated 123 amplicon products. Of these, two amplicons were shared by all the accessions whereas 112 were polymorphic at least in two pair wise comparison. Nine unique bands identified were as low as 0.32 kb M.W. to as high as 3.5 kb and were confined to eight collections. All primers produced polymorphic amplicons though the extent of polymorphism varied with each primer. The primer OPF-17 was found to be most powerful and efficient as it generated a total of 17 bands of which 15 were polymorphic. RAPD markers data were analysed statistically using NTSYSpc.2.02e software and a dendrogram was generated using Jaccard’s similarity coefficient. The similarity coefficient values varied from 0.19 to 0.91. Grouping analysis revealed the categorization of 99 germplasm into 12 major branches with different level of similarity. Three branches namely branch 2, 3 and 5 out of 12 had only one accession. Branch 1 which consisted of three accessions was the most divergent as revealed by Jaccard’s similarity coefficient. Branching pattern of the accessions did not show any correlation with morphological data or altitudinal alignment of the accessions.     

An Assessment of the Genetic Diversity within a Collection of Wild Cranberry (<Emphasis Type="Italic">Vaccinium macrocarpon</Emphasis> Ait.) Clones with RAPD-PCR

Forty-three wild cranberry (Vaccinium macrocarpon Ait.) clones collected from four Canadian provinces and five cranberry cultivars were assessed for genetic variability by using random amplified polymorphic DNA (RAPD)-PCR. Fourteen primers generated 161 polymorphic RAPD-PCR bands. A substantial degree of genetic diversity was found among the wild cranberry collections. Cluster analysis by the unweighted pair-group method with arithmetic averages (UPGMA) separated the wild clones and three cultivars into five main clusters, and identified the two remaining cultivars as outliers. Furthermore, within four clusters, the genotypes tended to form sub-clusters that were in agreement with the principal coordinate (PCO) analysis. Geographical distribution explained 10% of total variation as revealed by analysis of molecular variance (AMOVA). The RAPD markers detected a sufficient degree of polymorphism to differentiate among cranberry clones and cultivars, making this technology valuable for germplasm management and the more efficient choice of parents in current cranberry breeding programs.