利用AFLP鉴定柑橘变异

利用AFLP分析鉴定了7个柑橘变异类型,从64对引物组合中筛选了8对引物组合,共得到334个扩增位点2099条带数据。7个变异类型与亲本品种之间多态性带为36至92条,变异类型的多态性带数与亲本品种总带数的比例为0.1721至0.4182。说明变异类型的遗传基础已经发生了不同程度的改变,7个柑橘变异类型应是芽变,具备了成为新品种的遗传基础,特别是红皮冰糖脐橙选育成功将填补高糖脐橙空白。研究同时表明,AFLP具有扩增谱带多,谱带清晰,扩增信息量大的特点。根据谱带位点差异即可鉴定柑橘变异是否遗传,多态性位点数也反映了性状变异程度。AFLP方法1次可同时鉴定多个变异类型,是柑橘新品种选育早期鉴定灵敏、可靠、有效的方法。     

部分板栗品种遗传多样性的AFLP分析

利用荧光标记AFLP技术,采用7对M+3和E+3引物组合对30份板栗和日本栗栽培品种进行了总基因组DNA水平上的多态性检测,共获得962条可统计的条带,其中852条呈多态性,多态性带百分率达89％。揭示了板栗丰富的遗传多样性。7组引物在30个品种中检测到数目不等的品种特异带型,对供试板栗品种具有一定的鉴别价值。7对引物能将30个板栗和日本栗品种完全区分开。聚类分析结果表明,多数来源地相同的板栗品种资源表现出较为密切的亲缘关系。     

草莓品种亲缘关系的AFLP分子标记分析

 利用AFLP技术对我国国家草莓资源圃内保存的107个草莓品种进行了DNA多态性分析。从64对“2 + 2”引物中筛选出10对引物应用于扩增基因组DNA, 共获得清晰可辨的581条带, 其中412条为多态性带。UPGMA方法聚类分析结果显示, AFLP分子鉴定结果与其系谱关系非常一致, 表明AFLP指纹图谱分析能很好地鉴别草莓品种之间的遗传关系。     

萱草部分野生种和栽培品种亲缘关系的AFLP分析

 借助AFLP标记对35份萱草野生种和栽培品种进行亲缘关系研究, 结果表明, 7对引物组合对萱草共扩增出条带380条, 其中多态性条带357条, 平均多态性达到93.39% , 单对引物扩增条带19～85条, 平均每对引物组合扩增多态性条带51条。7对引物扩增出的多态性条带均超过90.0%。种质资源相似系数为0.3822～0.9656, 平均相似系数为0.7039。UPGMA聚类结果将供试材料分为3类, 即早花、中花和晚花类, 同一产地的品种基本能聚在一起。AFLP标记技术能较好地从分子水平揭示萱草种质资源的亲缘关系。     

杏品种的RAPD分析

实验从S系列(Sangon公司)100个引物中筛选出能稳定扩增的18个引物,用这些引物首次对新世纪与红丰等20个杏品种进行了扩增,均具有多态性。共扩增出218条带,106条具有多态性,多态性比例为48.6%;建立了红丰及新世纪等优新品种的DNA指纹图谱,找到了它们的特异谱带。利用特异谱带结合DNA指纹,可将参试品种鉴别出来。同时利用STATISTIC软件,根据RAPD标记相似系数采用UPGAM聚类策略,对20个杏品种进行了聚类分析。在LD=0.4时划等值线,将参试品种分为7类。     

用AFLP分子标记鉴定冬枣自然授粉实生后代杂种的研究

 应用AFLP技术对65株冬枣自然授粉实生后代进行杂种鉴定, 从81对EcoRⅠ和MseⅠ选择性引物中筛选出12对带型清晰且多态性较高的引物。用这12对引物共扩增出517条谱带, 其中多态性带376条, 多态性百分率为72.7%。共扩增出金丝小枣特征带10条, 鉴定出34株冬枣×金丝小枣杂交实生苗;扩增出尖枣特征带7条, 鉴定出冬枣×尖枣杂交实生苗15株; 不能确定的有16株。     

利用RAPD标记鉴定和区分梅花42个宫粉型品种

 利用RAPD标记，对形态特征相似的42个宫粉型梅花品种的基因组DNA进行随机引物PCR扩增。从120个引物中筛选出21个引物，共扩增出96条谱带，其中多态性谱带68条，占扩增谱带数的70．8％ ，品种问相似系数范围为0．737～0．961。在21个引物中，7个引物扩增的谱带可区分27个品种，7个引物相互组合可区分其余的l5个品种。梅花品种RAPD指纹图谱的建立可以为品种知识产权保护提供依据。     

辐射诱变和芽变柑橘品种(系)的AFLP分析

为了对广东省近年来新选育的16个60Co-γ辐射诱变和芽变柑橘优良品系(株系)进行遗传差异分析,应用AFLP技术,对3个亲本和16个辐射诱变和芽变的柑橘品种(系)进行了遗传差异分析。从64对引物中筛选出9对多态性高、分辨能力强的引物进行分析,结果表明:(1)与各自的亲本相比,供试16个辐射诱变和芽变的柑橘变异类型的遗传基础均已经发生了不同程度的变化。(2)利用引物EcoRⅠAAG+MseⅠCTT绘制供试样品的AFLP指纹图谱,并根据各自的差异带或特征带区分了19个柑橘试材。(3)对AFLP扩增结果进行聚类分析,根据相似系数0.77的水平可将19个样品分为4组。红江橙品系中的华青1号与亲本间的相似性系数仅为0.647 7,可将其单独划为一组。可为柑橘新品种选育研究的早期鉴定提供参考依据。     

基于SRAP的三倍体无子西瓜品种指纹分析和杂种纯度鉴定

为了解三倍体无子西瓜品种SRAP(Sequence-Related Amplified Polymorphism)扩增特点及其在三倍体西瓜杂种纯度鉴定中的适应性,利用SRAP引物组合对当前生产上推广的9个三倍体无子西瓜品种进行了指纹分析和杂种纯度鉴定研究。结果表明,1)30对引物组合中筛选出多态性引物组合24对,共产生109条多态性带,平均每对引物组合产生多态性带4.5条,单个组合的多态性比率在12.5%～80%。2)结合多对SRAP引物组合,可以鉴别不同品种。3)在黑蜜5号及其父母本的指纹分析中,发现有1对引物组合(ME4/EM3)在黑蜜5号杂交种上扩增出了特异的条带,可以区分母本和杂交种,试验证明该引物组合能够用于黑蜜5号的杂种纯度鉴定。     

不同梨品种的AFLP分子鉴别

利用8对引物对30个梨品种进行AFLP分析。结果表明:共检测到203个标记,其中多态性标记为135条,占所有标记数的66.5%,证明不同梨品种在DNA水平上存在广泛的遗传差异。根据AFLP标记结果计算不同梨品种间的遗传距离,通过聚类,发现所有品种聚为4个组:第1组主要是日本梨及与日本梨有亲缘关系的部分品种;第2组为中国砂梨品种;第3组只有2个品种:黄冠梨和清香梨,它们的一个共同亲本为新世纪梨;第4组是康德梨,其亲本中有1个是西洋梨,被单独聚为一类。从聚类结果看,不同的品种之间表现出了一定的地域相关性。     

An assessment of genetic variability and relationships within Asian pears based on AFLP (amplified fragment length polymorphism) markers

A total of 100 Pyrus L. accessions native mainly to East Asia were subjected to amplified fragment length polymorphism (AFLP) analysis to evaluate genetic variability and relationships among the accessions. Six AFLP primer combinations produced a total of 459 fragments, of which 410 were polymorphic with a polymorphism percentage of 89%. The Dice's similarity coefficient among pear accessions ranged from 0.671 (P. betulaefolia Bge and P. elaeagrifolia Pall.) to 0.947 (‘Umajirou’ and ‘Immuraaki’). Occidental pears generally had low similarities to Asian pears. The dendrogram generated from all the accessions by unweighted pair-group method of arithmetic analysis (UPGMA) cluster analysis clearly distinguished Occidental pears from accessions of East Asia. P. ussuriensis Maxim., P. betulaefolia and P. communis L. clustered separately into independent groups in accordance with their morphological classification. Japanese pear cultivars formed two groups with some Chinese white pears and Chinese sand pears. Chinese white pears and Chinese sand pears independently formed their own groups and also mingled into mixed groups in the dendrogram. Therefore, Chinese white pears were treated as a cultivated group or an ecotype of P. pyrifolia: P. pyrifolia White Pear Group. The information obtained from this study will be of great help for understanding the origin and evolution of Asian pear cultivars.     

Genetic relationships of gladiolus cultivars inferred from fluorescence based AFLP markers

Gladiolus is one of the important commercial flowers with a large number of cultivars. However, genetic relationships among its genotypes have not been reported. This study analyzed genetic relatedness of 54 gladiolus cultivars using amplified fragment length polymorphism (AFLP) markers. A total of 24 AFLP primer pairs with three samples were initially screened, from which 9 primer sets that showed clear scorable and highly polymorphic bands were selected for AFLP reactions. Fluorescence-labeled amplification products were subjected to electrophoresis and then analyzed using an automated sequencer. A dendrogram was constructed by the unweighted pair group method using the arithmetic average (UPGMA). The number of AFLP fragments generated per primer set ranged from 10 to 151 with fragment sizes varying from 50 to 450 bp. A total of 660 AFLP fragments were detected, of which 658 (99.70%) were polymorphic. All the primers except E-AGG/M-CTA displayed 100% polymorphism. All cultivars were clearly differentiated by their AFLP profiles. The AFLP data were compared with previously obtained RAPD data and combined to generate a common dendrogram. The first cluster was dominated with indigenously bred cultivars while the second was dominated with exotic cultivars. This shows that most of the exotic cultivars as well as indigenous cultivars are closely related with each other. However, two indigenous cultivars viz., Pusa Suhagin and Pusa Archana share genetic similarity with exotic cultivars. Among the genotypes selected for the investigation, Pusa Gunjan was identified as the most distinct genotype. The AFLP markers developed will help future Gladiolus cultivar identification, germplasm conservation and new cultivar development. The assessed genetic relationships among gladiolus cultivars may enhance the efficiency of breeding program by selecting desirable parents with reduced breeding cycle.     

甘肃中部梨种质资源的AFLP分析

采用AFLP分子标记技术对甘肃中部梨种质资源的遗传多样性进行分析。结果表明,6对AFLP引物在40份甘肃梨种质中共扩增出472条带,其中多态性带为404条,多态率高达86%,显示了甘肃中部梨资源丰富的遗传多样性。任何一对引物可以鉴别所有40份资源,显示了很高的鉴别能力。采用UPGMA聚类分析法构建的系统树在相似系数为0.72时将40份种质分为7个组:西洋梨、新疆梨、白梨(砂梨)、木梨、褐梨组和2个秋子梨组。所有白梨品种与唯一的砂梨品种黄花梨聚为一个大组。形态学上归属不明的品种分别聚类到西洋梨、白梨、木梨和秋子梨组中。研究表明基于AFLP标记的梨资源分类体系可以反映甘肃地方梨品种和类型间的遗传多样性和亲缘关系。     

部分桂花栽培品种的AFLP分析

 从22对AFLP引物中筛选出6对用来检测22个桂花品种和木犀属3个种的多态性位点,共检测到171个位点,其中多态性位点104个,占60.8%。利用DPSv 3.11软件计算25个样品之间的Nei遗传距离,并按照非加权算术平均数聚类方法（UPGMA）,构建树状分支图。AFLP分析结果表明：桂花花色较深的品种之间和花色较浅的品种之间分别存在着较近的亲缘关系,而花色深浅不同的两类品种之间亲缘关系较远。从聚类图上看,22个桂花品种中最后聚在一起的分别是3个银桂品种（遗传距离0.19处）和3个四季桂品种（遗传距离0.21处）,说明四季桂类和银桂类中的部分品种与金桂品种群和丹桂品种群有较远的亲缘关系。从分类上看,AFLP分析结果与传统的以形态特征为基础的分类结果并不完全一致。     

Molecular characterisation of Afghan pistachio accessions by amplified fragment length polymorphisms (AFLPs)

Summary

In Afghanistan, pistachio (Pistacia vera L.) is found mainly in the wild in natural forests. In this study, 17 wild Afghan pistachio accessions and three cultivated genotypes were characterised using amplified fragment length polymorphisms (AFLPs). Material was sampled mainly from the Kunduz and Takhar regions. A total of 288 AFLP fragments were generated using eight AFLP primer combinations. The number of amplified fragments varied from 18 – 48 per primer combination, and 136 bands were polymorphic, with an average of 17 polymorphic bands per primer combination. The percentages of polymorphic bands ranged from 26.2 – 79.2 per primer pair. According to UPGMA clustering of the Nei and Li distance matrix, the 17 wild accessions were grouped according to their region of origin and were distinct from the three cultivars. The AFLP technique was found to be effective for characterising wild P. vera material, which was genetically the closest to cultivated pistachio.     

Identification of 57 sweet orange cultivars using AFLP markers

Summary

Sweet orange (Citrus sinensis) represents an important group of Citrus fruit; however, the identification of sweet orange cultivars during vegetative growth can be difficult. A study on the genetic identification of sweet orange cultivars may be significant for the sweet orange nursery industry, for cultivar-rights protection, and is important for the genetic evaluation and conservation of these orange cultivars. In this study, amplified fragment length polymorphism (AFLP) markers were used to genotype 57 sweet orange cultivars. Ten PCR primer pairs generated 629 unique AFLP bands, with a size range of 50 ? 500 bp. Seventy-four bands (11.8%) were polymorphic. On average, each primer pair produced 62.9 fragments, with 7.4 polymorphic fragments. A dendrogram of the 57 sweet orange cultivars was constructed based on an UPGMA analysis using Jaccard?s coefficients of similarity. This provided a clear comparison of the genetic variation between cultivars and an ability to identify them. From Jaccard?s coefficients, 56 of the 57 cultivars examined were genetically close, with coefficient values ≥ 0.985. ?Variegated Navel? was less closely-related, with a much lower coefficient value (0.94). Among the 57 cultivars, 28 sub-groups, some consisting of only one cultivar, could be separated by their AFLP fingerprints. Compared to ISSR and SSR markers, AFLP seemed to be the preferential marker technique for the identification of sweet orange cultivars.     

枣树品种品系的AFLP分析

为建立完善枣种质资源分类鉴定技术体系,采用了基因组DNA-AFLP分子标记技术,从32对引物中筛选出了E-AAC/M-CCT、E-AAC/M-CAT、E-ACT/M-CCG、E-AAG/M-CCC和E-ACC/M-CAT等5个多态性好、分辨率高的引物组合,共扩增213条带,其中多态性带172条,多态性百分率76.16%。通过计算品种间单匹配系数,不加权算术平均值法绘制聚类图,可将28个供试品种和品系全部区分开;通过观察指纹图谱,也可明显看出新品系与其原种的差异带的多少,可快速准确地区分和确定新变异品系;另外,还可消除枣种质资源同名异物或同物异名现象。从而表明,AFLP技术是鉴定枣品种的有效方法。探讨了基因组DNA-AFLP分析技术的各关键步骤,并展望AFLP技术在枣品种鉴定上的应用前景。     

栽培番茄品种指纹图谱的AFLP分析

以62份栽培番茄为材料,利用65对AFLP引物进行选择性扩增,选出5对多态性丰富、带型清晰的引物进行扩增,共获得85个多态性位点,占总扩增位点的35.41%;选出清晰的16个位点构建了62份栽培番茄的AFLP指纹鉴别图谱。结果表明,由引物组合P55M48、P58M68、P21M49、P77M51、P58M60所构建的DNA指纹鉴别图谱可将供试材料完全区分开。     

苹果属观赏海棠实生单株亲本的AFLP鉴定

应用AFLP技术, 对以苹果属观赏海棠品种王族(Malus ‘Royalty’) 为母本, 以5个材料为父本的38个实生单株进行多态性分析。结果表明: 筛选出的9对分辨率高的引物共扩增出349条带, 其中多态性带303条, 多态位点百分率达86.8%。根据聚类分析和电泳图谱扩增得到亲本材料的特征带, 鉴定出19株实生单株的父本可能为‘宝石’ (Malus‘Jewelberry’) ; 7株实生单株的父本可能为海棠花(Malus spectabilis Borkh. ) ; 3株实生单株的父本可能为草莓果冻(Malus‘Strawberry Parfait’) ; 不能确定其父本的实生单株有9株。