杨树重要品种( 无性系) 的AFLP 指纹分析

品种的准确鉴定及其遗传相关性的了解对杨树育种和品种管理具有非常重要的意义。本试验采用AFLP对来自青杨组和黑杨组的21 个重要杨树品种( 无性系) 的鉴定与遗传相关性进行了研究。结果显示, 筛选的4对AFLP 引物总共产生了181 条多态性带,尤其是每对引物对每个品种都产生了独特的指纹图谱; 聚类分析和多维尺度分析将试验材料大体上分为五类, 结果不仅显示了组间不同品种的差异, 而且大体上区分了我国原生品种和外来品种。本研究表明, 所有品种都可被筛选的引物准确鉴定, 遗传相关性的推断结果与它们的系谱或分类基本一致。另外, 本研究还表明AFLP 技术完全可用于大规模地构建杨树树种DNA 指纹图谱、进行树种鉴定和遗传相关性的研究。     

吴茱萸的AFLP指纹图谱的初步研究

首次报道中草药植物吴茱萸基因组DNA指纹图谱的研究。吴茱萸是贵州省内经济价值极高的中草药之一,采用扩增片段长度多态性（AFLP）技术来分析来自不同地区的石虎、疏毛、大花吴茱萸3个品种的DNA指纹图谱,从18对引物中筛选出3对引物对19份材料的DNA检测,共得到93条带,其中多态性片段57条（平均61.3%）。3对引物组合从DNA指纹图谱上将19份材料完全区分开,结果表明AFLP技术是鉴别吴茱萸相近品种的有效方法,是形态学鉴定方法的有益补充;UPGMA方法聚类分析显示19份种质材料间的相似系数为0.235～0.941,表明吴茱萸种质间的遗传多样性丰富;余庆地区种植基地的石虎和疏毛样本聚为一类,提示人工栽培影响到吴茱萸的遗传特性。     

用AFLP分子标记探讨吴茱萸的遗传多样性

用AFLP标记分析研究吴茱萸与其变种石虎和疏毛吴茱萸之间的遗传多样性.采用AFLP技术,从18对引物中筛选出3对引物.对19株不同地域的3种吴茱萸AFLP指纹图谱进行了分析,计算不同品种间的遗传距离和构建吴茱萸的聚类分析树状图.3对引物共扩增出93条带,其中57条(61.3%)呈多态性,吴茱萸种质内遗传距离为0.059～0.765;在相似系数0.48的水平上.19份吴莱萸材料可以大致分为2个类群.吴茱萸不同品种问遗传距离差异较大,遗传分类在一定程度上与传统的分类方法是一致的,这暗示遗传变异与地域分布有相关性.     

云南栽培种及野生种芋头的AFLP指纹分析

目的:分析云南芋头种质资源遗传多样性。方法:应用扩增片段长度多态性(AFLP)指纹技术,用3对AFLP引物对采集自云南省的9份芋头栽培品种及1份野生品种进行研究,分离AFLP多态性条带。结果:共分离到60个AFLP多态性条带,AFLP多态位点百分率为96.77%,云南芋头种质资源在DNA分子水平上表现出丰富的遗传多样性。聚类分析将10份芋头品种分为2组,遗传距离为0.101～0.908。结论:AFLP指纹技术是筛选品种间差异基因的有效方法,研究结果为云南省芋头品种鉴定、遗传相关性分析、特殊功能基因的分离等工作提供了一定的理论基础。     

红麻微卫星DNA标记指纹图谱的构建

DNA指纹图谱对新品种选育、种质资源保存和管理具有重要的意义。然而,利用SSR标记构建红麻DNA指纹图谱的研究仍十分有限。在本研究中,利用课题组开发并筛选出的131对SSR引物,分析不同来源的96份红麻种质资源,包括红麻品种审定的区试对照品种福红952。结果表明,131对引物共扩增出375条带,平均每对引物扩增出2.6条带。以遗传相似系数0.614为切割线时,可以分为2个类群,52个为类群P1,44个为类群P2;以遗传相似系数0.710做切割线,可分为5个亚群。利用这131对引物标记所得的数据成功绘制了一份85个品种独特的指纹图谱,其中福红952可被HcEMS238引物特异识别。其他11份因存在遗传相似性高的现象,未被识别。上述结果为红麻品种的真实性鉴定及遗传多样性分析提供依据。     

水生实验动物剑尾鱼的DNA指纹图谱构建

目的剑尾鱼是由原良种委员会审定并由农业部公布的水生实验动物,在遗传学研究、水环境污染监测、细菌性疾病研究方面显示出较好的应用前景。为了对剑尾鱼选育系进行种质资源监测、区分选育系与非选育以及鉴定近交系纯度,本研究采用微卫星DNA进行水生实验动物剑尾鱼的指纹图谱构建。方法根据相关报道设计合成了50对微卫星引物,对几个剑尾鱼品系的种质资源进行检测,筛选品系间差异性引物;确立剑尾鱼核心引物,用EXCEL散点图绘制DNA指纹图谱模式图;并将数字化指纹数据输入珠江水产研究所鱼类种质鉴定软件V1.0,形成剑尾鱼标准化指纹图谱鉴定数据库。结果共获得剑尾鱼品系间特异标记5个可用于剑尾鱼近交系鉴定,确立46个微卫星标记为核心引物,构建剑尾鱼选育系RR-B系、RW-H系和非选育的野生品种的DNA指纹图谱。结论本研究筛选出的微卫星标记与构建的指纹图谱,可用于剑尾鱼3个品种间的品种鉴定、纯度检测及遗传监测。     

洋桔梗品种遗传多样性的AFLP分析

该研究以洋桔梗(Eustoma grandiflorum)2个品种‘玛丽艾基粉色’和‘圣剑白底紫边’为试材,提取叶片DNA,经过EcoRⅠ/MseⅠ双酶切、连接、预扩增、选择性扩增,建立了洋桔梗的AFLP最佳反应体系;并以64个常用引物组合进行扩增,得到154个多态性条带,从中筛选出扩增条带较多且多态性较好的4个引物组合(E-ACA/M-CTC,E-ACC/M-CAC,E-AGC/M-CTT,E-ACT/M-CAG),其多态位点百分率均值为24.36%。利用上述4个引物组合,以最佳反应体系为基础,构建了7个常见洋桔梗品种的AFLP指纹图谱,统计7个品种各4个引物组合在1 000~300bp区间7个区段的扩增条带,并将各个品种的AFLP指纹图谱转换成各品种4组7位数构成的28位特异数字指纹,极大地方便了种质比较及鉴定;7个品种间的遗传相似系数介于0.683 5~0.860 8之间,平均值为0.774 6。研究结果为进一步进行洋桔梗的种质研究及利用奠定了基础。     

莲品种DNA指纹图谱的构建

为了克服单纯依据形态特性鉴定品种的局限性, 我们开展了莲品种DNA指纹图谱构建研究, 旨在对其品种的快速准确鉴定及专利权保护等起一定作用。本研究以圆明园保存的72个莲品种为实验材料, 用来自不同地点的1,409份野生莲(Nelumbo nucifera)和58份美洲黄莲(N. lutea)群体样本作遗传背景参照。从104对核微卫星引物(nSSR)中筛选出15对, 从17对叶绿体微卫星(cpSSR)引物中筛选出2对, 共17对引物作为72个莲品种DNA指纹鉴定的条码。15对nSSR引物共检测到94个等位基因(平均6.27个), 其中11个属于美洲黄莲, 65个属于野生莲, 18个不能区分; 多态信息含量(PIC)介于0.3899-0.8023之间 (平均0.5748)。2对cpSSR引物共检测到13个单倍型, 其中9个属于野生莲, 4个属于美洲黄莲。全部17对引物标记结果显示, 共有19个品种含有美洲黄莲遗传组分, 其中8个母系来源于美洲黄莲; 有36个品种(涉及12对引物)具有至少1个特有基因型; 最少8对引物组合可完全区分开68个品种。有2组共4个品种组内全部17对引物均不能区分。本研究通过核心引物组合法使68个莲品种获得特异性DNA指纹。推荐13对nSSR和2对cpSSR共15对引物作为莲品种鉴定的核心条码, 并建议将形态特征与DNA指纹相结合作为莲品种的鉴定标准。     

鉴定烟草种质资源SSR核心引物筛选和验证

核心引物对遗传多样性分析、种质资源鉴定、品种纯度和真实性检测、指纹图谱构建等研究具有重要价值。本研究利用均匀分布于烟草24条染色体的278对SSR引物,对20份亲缘关系相对较远的烟草材料进行初步筛选,筛选出32对引物。随后再加上10份亲缘关系较近的材料（共30份）对引物进行复筛,最终确定14对为核心引物。将14对引物对39份烟草材料进行进行系谱分析、品种遗传多样性分析和农艺性状分析聚类,结果表明该套SSR核心引物适用于烟草种质资源鉴定和遗传多样性分析。     

中国南瓜海南农家品种间的遗传特异性分析和DNA指纹图谱构建

在收集中国南瓜海南农家品种的基础上,本研究应用ISSR和SRAP标记技术对28份海南农家品种间的遗传特异性进行了分析,并构建指纹图谱,为中国南瓜海南农家品种鉴定、评价、保护和利用提供科学依据。结果表明,所供试的品种间存在显著的遗传特异性,具有特殊的遗传基础或背景,所筛选的6个ISSR引物和11对SRAP引物共产生了10个特异标记和11条唯一缺失带;应用ISSR引物组合UBC807/UBC814/UBC844/UBC868和UBC808/UBC814/UBC844/UBC868,以及SRAP引物组合Me1/Em2 Me1/Em10 Me2/Em3和Me1/Em1 Me1/Em10 Me8/Em3分别绘制了四张28份中国南瓜海南农家品种的DNA指纹图谱,所构建的DNA指纹图谱直观、简单。ISSR标记和SRAP标记技术可有效应用于中国南瓜海南农家品种DNA指纹图谱的构建和遗传特异性鉴定。     

Genetic diversity in bambara groundnut (Vigna subterranea (L.) Verdc) landraces revealed by AFLP markers.

Bambara groundnut (Vigna subterranea (L.) Verdc), an African indigenous legume, is popular in most parts of Africa. The present study was undertaken to establish genetic relationships among 16 cultivated bambara groundnut landraces using fluorescence-based amplified fragment length polymorphism (AFLP) markers. Seven selective primer combinations generated 504 amplification products, ranging from 50 to 400 bp. Several landrace-specific products were identified that could be effectively used to produce landrace-specific markers for identification purposes. On average, each primer combination generated 72 amplified products that were detectable by an ABI Prism 310 DNA sequencer. The polymorphisms obtained ranged from 68.0 to 98.0%, with an average of 84.0%. The primer pairs M-ACA + P-GCC and M-ACA + P-GGA produced more polymorphic fragments than any other primer pairs and were better at differentiating landraces. The dendrogram generated by the UPGMA (unweighted pair-group method with arithmetic averaging) grouped 16 landraces into 3 clusters, mainly according to their place of collection or geographic origin. DipC1995 and Malawi5 were the most genetically related landraces. AFLP analysis provided sufficient polymorphism to determine the amount of genetic diversity and to establish genetic relationships in bambara groundnut landraces. The results will help in the formulation of marker-assisted breeding in bambara groundnut.     

A genetic linkage map of willow ( Salix viminalis) based on AFLP and microsatellite markers

The genus Salix (willow) contains a number of species of great value as biomass crops. Efforts to breed varieties with improved biomass yields and resistances to pests and diseases are limited by the lack of knowledge on the genetic basis of the traits. We have used AFLP and microsatellite markers to construct a genetic map of willow from a full-sib cross of the diploid species Salix viminalis (2n = 38). In accordance with a double pseudo-testcross approach, separate parental maps were constructed and merged to produce a consensus map comprising 291 AFLP and 39 willow microsatellite markers. Nineteen poplar microsatellites were also tested in willow. Five of these amplified loci, of which two were mapped. Linkage groups of the consensus map that could be identified in the parental maps are presented here and spanned 1,256.5 cM with an average interval between markers of 4.4 cM.     

基于EST-SSR标记的南方型黑杨主栽无性系鉴定及亲缘关系分析

以南方型黑杨引进后主栽区重点推广栽植的12个无性系为研究对象,利用EST-SSR分子标记进行多态性分析,并建立无性系鉴定的指纹图谱,为优良黑杨无性系的推广提供依据。用EST-trimmer对NCBI数据库中的EST序列进行分析、MISA软件找出SSR位点后运用primer3在线设计获得的30对EST-SSR引物,对12个黑杨无性系进行PCR扩增。经过筛选获得18对多态性引物,对12个黑杨无性系进行扩增获得88个等位基因,多态率为70.5%,平均观察等位基因数为2.1463,平均Shannon’s多样性指数为0.5927。通过5对引物组合(EU147、EU43、EU11、EU164和EU81)可以将12个黑杨无性系进行区分,并在此基础上构建指纹图谱。亲缘关系分析发现无性系间的遗传相似系数为0.6020~0.9040,平均值为0.7691;无性系间的遗传距离较近,与其均来源于美洲黑杨和欧美杨的相似遗传背景有关。总之,EST-SSR标记可以对南方型黑杨主栽无性系进行有效鉴定。     

Distinction between cold-sensitive and -tolerant jute by DNA polymorphisms

Jute is the principal coarse fiber for commercial production and use in Bangladesh. Therefore, the development of a high-yielding and environmental-stress tolerant jute variety would be beneficial for the agro economy of Bangladesh. Two molecular fingerprinting techniques, random-amplified polymorphic DNA (RAPD) and amplified-fragment length polymorphism (AFLP) were applied on six jute samples. Two of them were cold-sensitive varieties and the remaining four were cold-tolerant accessions. RAPD and AFLP fingerprints were employed to generate polymorphism between the cold-sensitive varieties and cold-tolerant accessions because of their simplicity, and also because there is no available sequence information on jute. RAPD data were obtained by using 30 arbitrary oligonucleotide primers. Five primers were found to give polymorphism between the varieties that were tested. AFLP fingerprints were generated using 25 combinations of selective-amplification primers. Eight primer combinations gave the best results with 93 polymorphic fragments, and they were able to discriminate the two cold-sensitive and four cold-tolerant jute populations. A cluster analysis, based on the RAPD and AFLP fingerprint data, showed the population-specific grouping of individuals. This information could be useful later in marker-aided selection between the cold-sensitive varieties and cold-tolerant jute accessions.     

A comparative assessment of molecular marker assays (AFLP, RAPD and SSR) for white yam (Dioscorea rotundata) germplasm characterization

Several DNA‐based marker systems are available for genetic fingerprinting of plants but information on their relative usefulness for yam germplasm characterisation is lacking. The efficiency of RAPD, AFLP and SSR markers for the assessment of genetic relationships, and for cultivar identification and discrimination among 45 West and Central African white yam cultivars belonging to 22 morphotypes/cultivar groups was investigated. Dendrograms were produced based on band pattern scores using the UPGMA method. Results showed that each of the three techniques could unequivocably identify each cultivar, but that techniques differed in the mean number of profiles generated per primer (or primer pair) per cultivar, referred to as genotype index (GI). The order of merit based on this criterion in this study was AFLPs (GI = 2.56), SSRs (GI = 0.39) and RAPDs (GI = 0.35). Yam genotypes classified in the same cultivar group based on morphology were often genetically different, emphasising the need for molecular fingerprinting in yam germplasm characterisation. AFLPs showed the highest efficiency in detecting polymorphism and revealed genetic relationships that most closely reflected morphological classification.     

Investigations of genetic variation between olive (Olea europaea L.) cultivars using arbitrarily primed polymerase chain reaction (AP-PCR)

Characterization and selection of olive clones for the production of olive oil is essential in Turkey because of its profitable exploitation. AP-PCR (Arbitrarily-Primed PCR) is a technique that can distinguish the genetic relationship among plant species and other organisms. In this study, AP-PCR approach was used in order to determine the genetic relationship of different six olive clones. The purity of DNA is one of the most important factors affecting the product of the AP-PCR method. In this respect, modified genomic DNA isolation procedure from Oleae europaea clones was developed so that this procedure can be used to obtain plant genomic DNA from diverse aromatic plants, which produce essential oils and secondary metabolites. By following the optimized AP-PCR amplification protocol, unique DNA fingerprint profiles for each olive clone were produced. AP-PCR-generated unique DNA fingerprint profiles can be used in the identification, distribution and diversity of various olive cultivars.     

DNA fingerprinting of Cryptosporidium parvum isolates using amplified fragment length polymorphism (AFLP)

The genetic variability of 10 Cryptosporidium parvum isolates of human and animal origin was investigated using amplified fragment length polymorphism (AFLP). Analysis of fluorescent dye-labeled amplified products was carried out using an ABI PRISMS 377 DNA sequencer and ABI PRISMS GeneScan software. One-hundred and twelve primer combinations were evaluated using a single C. parvum isolate. The patterns generated were highly reproducible. For subsequent study, a subset of 9 primer pairs that yielded 30-90 DNA fragments after the polymerase chain reaction, within the size range of 50-500 bp, was used to screen the 10 C. parvum isolates, including 7 bovine, 1 equine, and 2 of human origin. The animal isolates produced identical fingerprint patterns with every primer combination tested. Of the 2 human isolates tested, 1 of the isolates, passaged in calves, generated the same AFLP DNA banding patterns as the animal isolates, whereas the other isolate, obtained directly from human feces, produced unique patterns. Polymorphism, detected by comparison of the fingerprint patterns of the latter human isolate with the common pattern shared by all other isolates, ranged from 17 to 35% for the 9 primer pairs. The results show that AFLP is a useful method for differentiating C. parvum isolates into 2 distinct genotypes.     

漆树品种的AFLP分析及评价(简报)

漆树(Toxicodendron vernicifluum(stokes)F.A.Barkley)隶属于漆树科(Anacardiaceae)漆树属(Tox- icodendron)的落叶乔木,是我国重要的特用经济林木。漆树栽培与生漆使用在我国已有几千年的历史,在长期栽培过程中形成了许多农家品种,它们具有一定的形态特点,适应一定的生长环境,并具有产漆量高、生漆品质好等特性。     

Molecular genetic analysis of black poplar (Populus nigra L.) along Dutch rivers

The genetic structure of remaining black poplar ( Populus nigra ) trees on the banks of the Dutch Rhine branches was investigated using the AFLP technique. In total, 143 trees, including one P. deltoides and some P. x euramericana , were analysed using six AFLP primer combinations which generated 319 polymorphic bands. The AFLP patterns showed that some of the trees sampled as P. nigra were clearly different. These deviating patterns were also observed for the P. deltoides tree and all trees already identified as hybrid P. x euramericana . Hybrids between the two species are morphologically sometimes difficult to distinguish from the species itself. Two important possible source populations for recolonization of the riverbanks of the river Rhine, consisting of mature flowering P. nigra trees, appeared to consist of only a few genotypes each. In contrast, young black poplar trees growing alone or in small groups downstream of the possible source populations appeared to be predominantly generatively derived because no clones of mature trees were found among them. Therefore vegetative propagation seems a very local strategy whereas colonization of new areas appears to occur through generative propagation. Whether the genetic diversity within these black poplars is sufficient for recolonization of river banks and survival of the metapopulation is a question for further research.