DNA分子标记在番茄遗传育种研究中的应用

本文综述了DNA分子标记在番茄遗传图谱构建、番茄种质资源研究与品种纯度的鉴定、番茄基因分子标记研究及番茄基因图位克隆方面的应用研究进展。Abstract:This paper reviewed the recent advance of the application of DNA molecular marker in various aspects of tomato breeding including genetic map construction,germplasm research and purity control of cultivars,identification markers linked to important genes and map-based gene cloning.     

DNA分子标记在果树遗传学研究上的应用

本文综述了近年来DNA分子标记在果树种质资源研究、分子遗传图谱构建、基因标记、辅助选择育种等方面研究的应用。Abstract:This paper reviewed the recent progress of the application of DNA molecular markers in various aspects of fruit tree genetics including germplasm reseach,genetic mapping,genetic tagging,marker asisted selection.     

偏分离分子标记的作图方法

对取自MAPMAKER软件小鼠F2群体(含333个体)的5个RFLP连锁标记数据作了共显性分子标记偏分离的分析。先确定选择类型的方程组(配子或合子),随后采用Newton-Raphson迭代法估算标记间的重组值。在构建分子标记遗传图谱时,如果两个相邻标记均存在偏分离,最好采用纳入偏分离因子的估算方法。在估计F2群体标记间偏分离重组距离上,用连续χ2检测方法比传统χ2检测更为准确。Abstract The comparative analysis of segregation distortions of the codominant markers data presented in software MAPMAKER are made, where five RFLPs markers involve in a mouse F2 population with 333 individuals. The successive χ2 test begins with the determinations of gametic or zygotic selection types, followed by the estimation of recombination fractions between two markers with the Newton-Raphson iteration method. It is better to use the molecular marker showing segregation distortion for constructing a genetic map, in the case of seriously skew segregation between both the adjoining markers. The successive χ2 test provides better accuracy than that of classical χ2 test for the estimation of the recombination values in F2 population with segregation distortion.     

单核苷酸多态性在作物遗传及改良中的应用

单核苷酸多态性（single nucleotide polymorphism,SNP）是等位基因间序列差异最为普遍的类型,可作为一种高通量的遗传标记。已建立了PCR扩增目标序列及其产物测序和电子SNP（eSNP）等多种发现和检测SNP的方法。玉米和大豆等作物也已开展了SNP分析。一些栽培作物种质的多样性不断减少,其结果使连锁不平衡（linkage disequilibrium,LD）增加,这有利于目的基因座上SNP单元型（haplotype）与表型的相关性分析。SNP已在作物基因作图及其整合、分子标记辅助育种和功能基因组学等领域展示了广泛的应用价值。Abstract:Single nucleotide polymorphism(SNP) is the most common type of sequence difference between alleles,which can be used as a kind of high-throughput genetic marker.Several different routes have been developed to discover and identify SNP.These include the direct sequencing of PCR amplicons,electronic SNP(eSNP) and so on.SNP assays have been made in many crop species such as maize and soybean.The elite germplasm of some crops have been narrowed in genetic diversity,increasing the amount of linkage disequilibrium(LD) present and facilitating the association of SNP haplotypes at candidate gene loci with phenotypes.SNP analysis has been broadly used in the field of plant gene mapping,integration of genetic and physical maps,DNA marker-assisted breeding and functional genomics.     

相关序列扩增多态性(SRAP)标记及其应用研究进展

SRAP是一项基于PCR技术的分子标记技术,利用其独特的引物设计对基因组的开放阅读框(ORFs)进行特异扩增,利用个体以及物种的内含子、启动子和间隔序列的不同,产生基于内含子和外显子的SRAP多态性。阐述了SRAP的原理和流程,详细论述了SRAP标记目前在植物遗传多样性、作物品种鉴定、遗传图谱构建等方面的研究进展及应用前景。     

PCR扩增近交系大鼠微卫星位点DNA多态性的研究

本实验选取大鼠7条染色体上的微卫星位点合成了10对引物,利用聚合酶链反应(PCR)扩增技术对国内北京和哈尔滨等4家单位提供的6个品系(SHR、SHRSP、LEW、RCS、WKY和F344)的8个近交系大鼠群体进行了DNA多态性分析的研究.结果表明:9个微卫星位点具有显著多态性;不同品系个体之间具有多态性;同一群体不同个体之间除SHR(哈)的SMST位点和WKY(哈)的AGT位点出现一定的差异外,其他均没有差异;不同地区同一品系的不同个体之间也存在一定的差异.该方法能有效地对近交系与杂交系、品系与品系、品系与亚系加以区分.因此,本实验为开展近交系大鼠遗传作图、基因定位和为实验动物的遗传背景监测提供可靠的信息,为大鼠遗传基因的研究提供了一个快捷简便、特异准确的方法。Abstract:In the experiment,It were selected that 20 primers were assigned on 7 chromosomes of inbred rat.DNA polymorphism of 8 colonies from 6 inbred rat strains(SHR、SHRSP、WKY、LEW、RCS、F344)were studied in national Beijing and Harbin using PCR-analyzed microsatellites.The results indicated that there were remarkable polymorphism in 9 microsatellite loci;There is a polymorphism among the various rat strains;and no polymorphism in the same rat strains except SMST locus of SHR(Harbin)and AGT locus of WKY(Harbin)and there is a little difference among the same inbred rat strains in different areas.The method of PCR-analyzed microsatellites can be used for distinguishing between inbred and outbred、different strains、strain and substrain.and it provides a lot of information for genetic mapping,gene location and heredity probe of the inbred rat strains,and a speedy、convenient method for genetic research of the inbred rat.     

DNA多态性分析技术研究进展

随着现代分子生物学的迅速发展,DNA多态性技术在很多领域得到了应用,本文简单总结了DNA多态性分析技术的发展,并展望了DNA多态性技术的应用前景。     

小麦叶片直接用于PCR和RAPD反应的方法

本研究以经过碱处理的小麦叶片直接作为PCR和RAPD反应的模板,并应用于小麦转基因目标性状的跟踪检测、品种多态性分析等方面。该方法具有快速、简便等特点,尤其是在受测群体较大时,此种方法更显得经济有效。Abstract:In this paper we introduce a new method of PCR and RAPD reaction by using a small quantity of wheat leaf tissue with alkali treatment.It can be used either to detect target gene or to analyze polymorphism of wheat varieties.This method is simple and reliable,especially for a large-scale detection.     

AFLP分析中多态性扩增产物的回收、克隆及鉴定

本研究在摸索和优化了水稻AFLP分析体系的基础上,发展了多态性AFLP产物的高效克隆方法。特异AFLP扩增产物直接从变性聚丙烯酰胺凝胶上分离纯化,再经过一至二轮PCR扩增,即可高效地克隆于pGEM-Teasy vector系统中。本实验利用该方法成功地克隆了水稻温敏核不育等位突变系546 0S和5460F间的4个多态性AFLP产物,Southern bloting分析证明其中3个产物在水稻基因组中为单拷贝序列,另一个为低拷贝序列。AFLP技术强有力的多态性检出能力再结合多态性扩增产物的高效克隆方法,为寻找与目标基因紧密连锁的分子标记提供了有力工具。Abstracts:An efficient method for cloning DNA fragment from denaturing polyacrylamide gels was developed to allow the isolation of specific bands obtained from amplified fragment length polymorphism(AFLP)products.After isolation and purification from the thin denaturing polyacrylamide gels,specific AFLP products were successfully cloned after one or two rounds of PCR reamplification.Using this method 4 polymorphic AFLP products between a pair of rice allelic lines differing for thermo-sensitive genic male sterile(TGMS)ene were cloned and it was confirmed that 3 of the AFLP products represented single copy sequences and the other 1 represented low copy sequence in rice genome.     

水稻一多拷贝微卫星DNA多态性分析

用一个多拷贝微卫星 DNA标记分析了238份栽培水稻的遗传多样性及遗传多样性从农家品种到现代栽培品种的动态变化。共检测出16种长度变异类型和32种表现型,但长度变异类型数、表现型数和表现型多样性水平在农家品种和现代栽培品种间没有显著差别。该标记在遗传资源评价和DNA指纹图谱建设中有重要的应用价值,也是研究环境或人工对多拷贝基因选择作用的一个良好模型。Abstract:The genetic diversity of 238 cultivated rice and its difference between landraces and modern cultivars have been assayed using a multiple copy microsatellite DNA marker.Although 16 length variants and 32 phenotypes were found in all accessions,no significant differences in number of length variants,number of phenotypes and phenotypic diversity were detected between landraces and modern cultivars.This marker is of great value in evaluating genetic germplasm and constructing DNA fingerprints of rice and provides an elite model for tracing effectiveness of natural and artificial selection on multiple copy genes.     

大麦6H染色体特异性标记的筛选和鉴定

从大麦、小麦和小麦－大麦６Ｈ染色体附加系ＲＡＰＤ分析筛选出对６Ｈ染色体特异的２个ＲＡＰＤ标记,转换为特异性ＰＣＲ标记,利用标记对不同植物材料进行ＰＣＲ扩增鉴定。表明凡含有大麦６Ｈ染色体的材料（Ｂｅｔｚｅｓ、Ｉｇｒｉ、ＣＳ６Ｈ附加系）均能扩增出特异带;而不含６Ｈ染色体的材料,包括小科、黑麦、长穗偃麦草、中间偃麦草、簇毛麦以及含有其他大麦染色体的小麦附加系均不主增出特异带。可见,２对ＰＣＲ引物具有大麦     

赤眼蜂分子鉴定研究进展

赤眼蜂是害虫生物防治中的重要天敌资源.该属种类繁多,已报道有200余种,其蜂种的正确鉴定与选择是影响其田间防效的重要因素.依赖雄成蜂外生殖器形态特征的传统赤眼蜂分类鉴定技术不仅对专业技术要求高、耗时费力,而且无法用于孤雌产雌品系的种类鉴定以及近缘种的区分.分子鉴定技术可以通过选择合适的分子标记,为赤眼蜂鉴定提供准确、便...     

植物染色体显微切割技术及其应用(综述)

本文综述植物染色体微切割、微克隆技术的原理和方法,以及该技术在植物学研究上的应用。     

A novel approach for the identification of bacterial taxa-specific molecular markers

AIMS: To develop and establish a methodology for an oriented and fast identification of species taxa-specific molecular markers useful for the identification of micro-organisms. METHODS AND RESULTS: From the complete microbial genomes available in Pfam database, taxa-specific protein domains were identified which lead to the selection of taxa-specific loci. This strategy was used to identify six genetic markers: four specific for Pseudomonas syringae pv. tomato, one specific for P. syringae pv. syringae and one specific for P. putida. The discriminatory potential of these loci was evaluated by Southern hybridization using several pseudomonad species and pathovars, by dot-blot hybridization and by multiplex PCR optimized for the simultaneous detection of P. putida, P. syringae pv. syringae and P. syringae pv. tomato. Sensitivity assays indicated a detection limit of approximately 10 pg of chromosomal DNA template needed for each bacterium. CONCLUSIONS: The proposed methodology was efficient on the selection of six Pseudomonas-specific markers able to discriminate Pseudomonas at the species and pathovar level. SIGNIFICANCE AND IMPACT OF THE STUDY: The oriented search of taxa-specific molecular probes described in this work, which can be easily extended to other groups of bacteria, will improve the accuracy and expedite the identification of micro-organisms by DNA-based molecular methods.     

小麦族中含St染色体组物种的特异分子标记的建立

以拟鹅观草(Pseudoroegneria spicata)、偏凸山羊草(Aegilops ventricosa)、二倍体簇毛麦(Dasypyrum villosum)、荆州黑麦(Secale cereale cv. Jingzhou rye)、普通小麦中国春(Chinese Spring)等15个物种为材料, 用200条10碱基随机引物进行RAPD分析, 筛选到拟鹅观草基因组中1个542 bp的特异DNA片段(GenBank登录号为DQ992032), 命名为OPH11542。根据OPH11542设计特异引物, 对小麦族物种进行PCR扩增, 发现拟鹅观草可以扩增出OPH11542以及分子量分别为742 bp (GenBank登录号为DQ992033, 记为OPH11742)和743 bp (GenBank登录号为EF014218, 记为OPH11743)的DNA片段, 而其他材料均未扩增出这3个片段。经序列比对结合多个软件的分析结果认为该3个片段为同一类新重复序列。利用特异引物对15份含St染色体的物种进行扩增, 发现含StY染色体组的物种均能扩增出OPH11742或OPH11743, 而含StH染色体组的物种均能扩增出OPH11542。这表明St染色体组在与其它染色体组组合形成多倍体的过程中往往会出现不同程度的重组或修饰。OPH11542、OPH11742和OPH11743可以作为检测St染色体的分子标记。     

大麦基因组中的微卫星标记及其应用

微卫星是以少数几个核苷酸为单位多次串联重复的DNA序列,是一种简单序列重复(simple sequence repeats,SSR),两侧一般是保守序列。由于它具有多态性高、共显性、容易用PCR检测和结果稳定可靠等特点,因此是一种十分理想的分子标记。大麦的微卫星DNA随机分布于基因组中,平均每一个微卫星基因座有3～18个等位基因,最高可达37个。SSR标记已广泛用于分子遗传图谱的构建、遗传多样性研究、种质鉴定、主要性状基因的定位及分子标记辅助选择育种等。大多数SSR标记集中在着丝粒附近区域,1HL、5HL和6HS明显缺乏SSR标记。大麦的SSR标记还有待进一步的开发。Microsatellite Markers and Applications in the Barley GenomeFENG Zong-yun1,2,3,ZHANG Yi-zheng1,LING Hong-qing31.College of Life Sciences,Sichuan University,Chengdu 610065,China;2.College of Agronomy,Sichuan Agricultural University,Ya'an 625014,China;3.The State Key Laboratory of Plant Cell & Chromosome Engineering,Institute of Genetics & Developmental Biology,Chinese Academy of Sciences,Beijing 100101,ChinaAbstract:Microsatellites,also called simple sequence repeats (SSR),are simple,tandemly repeated DNA sequences with a repeat length of a few base pairs,and are very ideally used as molecular markers because of their abundance,high level of polymorphism,co-dominance and ease of assay with the polymerase chain reaction (PCR) by selecting primers as the conserved DNA sequences flanking the SSRs,as well as better stability.The experiments showed that SSRs are randomly distributed throughout the barley genome,and there are 3~18 alleles at a single SSR locus,up to 37 alleles/locus.SSR markers have being widely applied in the construction of molecular genetic map,the study of genetic diversity,the identification of germplasm,gene mapping for important traits and molecular marker-assisted selection.Meanwhile,most of markers are strongly clustered around the centromeric regions of all seven linkage groups.As a result of the clustering,genome coverage with SSRs remains incomplete with an obvious lack of markers on the long arms of chromosomes 1H and 5H and short arm of chromosome 6H.Therefore,it is very potential and necessary to further develop SSR markers in barley.Key words：barley;microsatellite marker;simple sequence repeats;genetic diversity;molecular mapping     

Conversion of an RAPD marker to an STS marker for barley variety identification

Barley (Hordeum vulgare L.) variety identification is important to the malting and brewing industries. Because many new malting cultivars (varieties) are closely related, new and more effective identification techniques are needed. We report on a series of techniques used to convert an RAPD marker to a more stable STS marker that can identify barley Stander from Robust, an important distinction for the American malting and brewing industries. The techniques included DNA extraction, RAPD amplification, random cloning of all amplified fragments, selection of clones by insert size, DNA sequencing of select inserts, design of a barley-based primer pair, and detection of a single nucleotide polymorphism using restriction endonucleaseAlu I. The barley-based primer pair was used to further sequence the RAPD fragment. Five single nucleotide polymorphisms between Robust and Stander exist, one of which was detected by electrophoresing DNA fragments differentially restricted byAlu I. The conversion technique was different from ones previously reported in that it did not require manual extraction of DNA fragments from a gel. This could be applied to other situations in which RAPD marker conversion would be desirable.