番茄分子标记开发进展

分子标记辅助选择为选育由多基因控制的或易受环境影响的性状提供了一条有效的途径.番茄分子标记的开发工作已经进行了三十多年,到目前为止已有2 300多个不同类型的分子标记被开发和定位到12条染色体上.很多文献对这些分子标记的特点及其在番茄遗传育种中的应用进行了综述,但对其开发过程却很少提及.因此本文就主要类型的番茄分子标记的开发进展进行概括并分析其开发前景.     

ISSR分子标记及其在植物研究中的应用

ISSR(inter simple sequence repeat)分子标记是一种以微卫星序列为引物,进行多位点PCR扩增的技术.ISSR技术简易、快捷,同时兼备AFLP(扩增酶切片段多态性)、SSR(简单序列重复)和RAPD(DNA随机扩增多态性)等分子标记方法的优点.引物设计无需预知基因组序列,只要是目标区域的长度在可扩增范围内,就能扩增出微卫星重复序列间的DNA片段.ISSR标记以其高多态性,已被广泛应用于种质收集、品种鉴定、遗传多样性、系截系、遗传作图、基因定位、标记辅助选择、预测基因组SSR基序的丰度以及SSR引物开发等研究.本文对ISSR技术及其在植物研究中的应用进行全面的概述.     

iPBS分子标记研究进展

DNA分子标记是一种可以直接反映生物个体DNA水平差异的优良技术,自开发以来,在基因组作图和基因定位研究、基于图谱克隆基因、物种亲缘关系和系统分类中的应用等各个领域应用广泛。近年来,随着分子技术在应用过程中的不断完善、更新和发展,不同类型的DNA分子标记层出不穷。本综述基于反转录转座子的新型分子标记i PBS技术,介绍了iPBS分子标记的开发和基本的PCR反应体系,以及其在种质资源鉴定、遗传多样性研究、DNA指纹图谱构建、植物LTR类反转录转座子的序列分离等方面的应用和最新研究进展,以期为植物分子生物学研究及植物育种和改良提供参考。     

分子标记在松树遗传与进化研究中的应用

松树是世界上分布最广且最具经济价值的树种之一。近年来随着DNA分子标记技术的发展,分子标记已经广泛应用于松树的遗传多样性、亲缘关系、遗传图谱构建、基因定位及分子标记辅助选择等方向。本文从遗传多样性、亲缘关系、遗传图谱构建、基因定位及标记辅助选择等角度,评述了松树遗传与进化上常用的DNA分子标记如RFLP、RAPD、AFLP和SSR等的应用情况和进展。在遗传多样性方面,DNA分子标记技术已成为鉴定松树种间、种内遗传多样性的有效手段,并用于指导杂交育种;在亲缘关系判定方面,可以在分子水平上阐明生物系统演化及分类情况,揭示育种亲本的选配和种质资源的有效利用;在松树遗传图谱构建和基因定位及标记辅助选择方面也取得了显著进展,构建了二十多张连锁图谱,并对其生长、材性和抗性等性状进行了基因定位,获得了一些重要的与抗病相连锁标记。     

DNA分子标记在洋葱遗传育种研究中的应用

DNA分子标记技术的出现大大提高了遗传分析的准确性和选育品种的有效性,在植物遗传育种领域越来越受到重视.洋葱属于二年生植物,一个世代需要三年,具有育种年限长的问题.利用分子标记辅助选择是缩短洋葱育种年限,加快育种进程极其有效的途径.本文综述了DNA分子标记技术在洋葱遗传图谱构建、遗传多样性分析、种质鉴定、鉴定洋葱细胞质类型、重要性状的分子标记辅助选择和数量性状的基因定位等方面的应用,讨论了洋葱遗传育种中分子标记技术的现状及存在的问题,并对洋葱分子标记辅助育种的应用前景进行了展望.指出洋葱高密度的遗传图谱还有待于建成,对于数量性状标记的鉴定,还有待于进一步开发、快速和准确定位的QTL,关于洋葱抗病基因的标记需要深入研究.     

基于RNA-seq牡丹SSR标记开发及通用性分析

从牡丹花色候选基因中开发一套SSR标记,为牡丹花色分子遗传学研究和分子标记辅助育种提供帮助。本研究基于高通量转录组测序技术(RNA-seq)获得了278条涉及调控牡丹花色形成的Unigenes序列,利用SSRIT在128条Unigenes中检测到215个SSR位点,出现频率为46.04%。其中优势重复基序为二核苷酸、三核苷酸和六核苷酸重复,分别占总SSR位点的18.60%、41.86%和36.28%,优势重复基元为TC/GA和AT/TA,分别占二核苷酸重复的30.00%和27.50%。在此基础上,从中选择100对SSR引物合成,以牡丹品种基因组DNA为模板,验证其有效性和多态性。结果表明,有效引物50对(占50%),多态性引物12对(占24%)。12对多态性引物在芍药属12个不同种质内进行通用性检测,转移率范围为83.33%~100%,平均转移率为96.53%,表明牡丹SSR标记在芍药属内具有较高的通用性。利用高通量RNA-seq开发牡丹候选基因SSR标记可为牡丹功能基因挖掘、品种分子身份证构建、遗传多样性分析和分子标记辅助选择育种等提供重要的遗传资源。     

目标起始密码子多态性(SCoT):一种基于翻译起始位点的目的基因标记新技术

随着功能基因组学的发展,分子标记领域已开始从传统的随机或匿名性分子标记转向目的基因标记和功能性标记.目标起始密码子多态性(start codon targeted polymorphism,SCOT)标记是一种基于翻译起始位点的目的基因标记新技术,具有操作简单、重复性好等特点,根据ATG翻译起始位点侧翼保守序列来设计单引物,扩增产生偏向候选功能基因区显性多态性标记,适合不同层次实验室的不同领域的广泛应用,SCoT标记已开始在水稻和花生上得到运用.本文旨在介绍给研究者一种可以作为传统的RAPD、ISSR标记有效补充的目的基因标记新技术,希望其广泛的应用于生物多样性分析、遗传图谱构建、重要性状标记等方面.     

基于甘蔗热带种(LA-purple)全基因组序列的SSR开发及特征分析

现代甘蔗栽培品种(2n = 100~130)是由甘蔗热带种(2n = 80)与割手密(2n = 40~128)种间杂交而来, 形成异源多倍体、非整倍体作物, 使得甘蔗栽培品种中80%~90%的染色体来源于热带种。开发热带种基因组SSR分子标记, 有助于甘蔗遗传多样性分析、分子标记辅助选择、遗传图谱的构建等。本研究基于热带种LA-purple的全基因组测序数据的255 398个预测基因序列(累计总长为1 029 222 285 bp), 利用Perl程序与生物信息学软件结合, 发掘SSR位点, 获得了153 150个SSR位点, 平均每1.67个基因有1个SSR位点, 其中二、三核苷酸重复基序分别为39 556个和50 072个, 占总SSR位点数的58.5%。在二核苷酸重复基序中, TA/AT所占比例最高, 占41.4%, CG/GC所占比例最低, 占4.6%; 在三核苷酸碱基重复基序中, TGT/ACA所占比例最高, 为15.6%。在TA/AT重复类型中选取100个基序重复次数在60~90之间的SSR位点, 进行引物设计与合成, 在12个甘蔗属材料中进行PCR扩增分析, 从中筛选出52对具有多态性SSR引物, 其中有27对引物在研究的2个甘蔗栽培品种间表现为多态。这些基因组SSR标记的开发, 不仅可以用于甘蔗栽培品种DNA指纹图谱分析, 而且为甘蔗属不同种的遗传图谱构建、遗传多样性分析和重要性状的遗传机制解析奠定基础, 为甘蔗分子育种研究提供重要支撑。     

遗传标记在石榴种质资源研究中的应用

石榴属石榴科石榴属植物,具有较高的营养价值、药用价值和观赏价值。笔者从形态、细胞、生化和DNA分子等不同层面对遗传标记在石榴种质资源研究中的应用进行阐述,并对遗传标记方法的优缺点及存在的问题进行分析和探讨,最后提出了一些建议。遗传标记的应用,特别是分子标记的发展,将为石榴遗传多样性研究、亲缘关系分析、分类研究、品种鉴定、遗传图谱构建以及功能基因定位等相关研究开辟新的途径。     

ISSR分子标记技术在植物种质资源研究中的应用

ISSR是在SSR基础上发展起来的一种分子标记技术,兼具SSR、RAPD、RFLP、AFLP等分子标记的优点.本文综述了ISSR分子标记技术在种质资源鉴定和指纹图谱构建、遗传多样性和亲缘关系、基因定位和分子标记辅助选择等方面的研究进展.     

Genetic diversity of new maize hybrids based on SSR markers as compared with other molecular and biochemical markers

Genetic diversity of four new yellow single crosses, five new yellow three-way crosses, and five yellow inbred lines of maize (Zea mays L.) was studied using different molecular (SSR, ISSR, and RAPD) and biochemical markers (seed storage protein content). All markers were able to clearly separate the inbred lines in one cluster from the different types of hybrids. The correlation among the different types of molecular markers was moderately high according to the Mantel’s test (e.g. 0.67 between SSR and ISSR, 0.42 between SSR and RAPD, and 0.65 between ISSR and RAPD), indicating that the three techniques are efficient for evaluating genetic diversity in the maize genotypes. The correlation of biochemical markers (seed storage protein SDS-PAGE) with SSR, ISSR, and RAPD markers was 0.61, 0.74, and 0.48, respectively. It can be concluded that both molecular and biochemical markers are efficient to study the genetic diversity in maize. Among the different types of molecular markers, SSR is a more accurate marker-type because of its co-dominance and stability of results. It can also be said that biochemical and molecular markers are positively correlated and the correlation ranged between moderate to high. This could suggest using both marker types, separately or together, for genetic diversity studies in maize.     

Genetic diversity assessment and genotype identification in sugarcane based on DNA markers and morphological traits

Sugarcane is known for its highly complex genetics and more knowledge is needed for better use and conservation of genetic materials. In order to identify genotypes and to assess genetic diversity, diverse data sets such as morphological and molecular markers are used as a general approach. To evaluate the usefulness of different markers, important sugarcane genotypes in Argentina were characterized by AFLP, SSR and morphological traits. All genotypes characterized were grouped in one main cluster in dendrograms using two independent softwares. Interestingly, local genotypes grouped together with USA varieties and no clear genetic differentiation could be found probably due to intensive germplasm exchange between these breeding programs. The molecular markers tested were useful for genetic diversity assessment as well as for genotype identification. These markers should be included in the internationally established characters for sugarcane variety protection as they give a better view on whole genome complexity. Additionally, genetic similarities obtained from molecular markers will provide more accurate information to breeders than the pedigree method, especially when considering the asymmetric genetic inheritance of sugarcane. Morphological traits are valuable tools to identify genotypes since they reflect external resemblance more than genetic relatedness. When they were combined with molecular markers the dendogram obtained revealed genetic relationships and the genetic diversity was better estimated. In summary, both methods appear to be useful, complementing each other and should be used together to assist sugarcane breeders in estimating genetic diversity, electing parents for crossings, identifying superior lines and to protect intellectual property rights.     

Intron Polymorphisms: Species-specific and Interspecies-comparable Markers

As a component of genes, introns are much more variable than exons. Therefore, introns are an important source of genetic polymorphisms and can be used as genetic markers.     

Evaluating genetic variation and relationships among two bromegrass species and their hybrid using RAPD and AFLP markers

The two most widely grown bromegrass species in North America are smooth bromegrass (Bromus inermis Leyss.), a hay type grass, and meadow bromegrass (Bromus riparius Rehmann), a pasture type grass. Recently a hybrid bromegrass population between the two species has been produced as a dual-purpose hay-pasture grass. Molecular markers have the potential to improve selection procedures to enhance bromegrass breeding. The objective of this study was to use RAPD and AFLP markers to determine genetic relationships and variations among bromegrass populations. Forty-three RAPD markers from 21 primers and 83 AFLP markers from seven primer combinations were used. Both marker types were able to group the individuals into their respective populations. The relationships among the individuals within each of the populations were not similar between the two marker types. Analysis of molecular variance (AMOVA) detected greater within-population variation than among-population variation for both marker types. The highest variation was observed in the hybrid population followed by meadow and then smooth bromegrass. The inter-population distance from both markers indicated that the highest genetic distance was between meadow and smooth bromegrass and lowest between smooth and hybrid bromegrass, which reflect the breeding history of the hybrid population. This study showed that both markers are capable of differentiating bromegrass genotypes into their respective populations, detecting genetic variation and relationships of the populations. Results of this study suggest that these two markers can be used in the future to enhance the current breeding practices in bromegrass, however, AFLP markers would be the marker of choice due to the high number of polymorphic markers. This revised version was published online in July 2006 with corrections to the Cover Date.     

桉树的分子标记技术及遗传图谱构建进展

桉树是世界上的三大类速生树种之一，具有重要经济价值。本文着重论述了RFLP、RAPD、AFLP和SSR等4种DNA分子标记技术在桉树上的应用和推广，讨论了基于上述分子标记构建的桉树遗传连锁图的现状及其应用，提出了当前桉树遗传图谱构建存在的问题，进一步探讨了桉树的分子标记技术和遗传图谱构建的发展趋势。     

Development and validation of a PCR-based functional marker system for the brown planthopper resistance gene Bph14 in rice

Brown planthopper (BPH) is the most damaging rice pest globally. Resistant varieties are the most effective and environmental strategy for protecting the rice crop from BPH. Functional markers (FMs) designed from polymorphic sites within gene sequences affecting phenotypic variation are highly efficient when used for marker assisted selection (MAS). Bph14 is the first and only cloned insect resistance gene so far in rice. Compared to the sequences of its non-effective alleles there are a number SNP differences. In this study, the method of allele-specific amplification (ASA) was adopted to design a simple, co-dominant, functional marker Bph14P/N for Bph14. Bph14P/N was combined with two specific dominant markers: one, named Bph14P, targets the promoter region of Bph14 and amplifies 566 bp fragments; and the other, Bph14N, targets the LRR region of bph14 and amplifies 345 bp fragments. Specificity and applicability of the functional marker system were verified in two breeding populations and a Chinese mini core collection of Oryza sativa. We recommend the use of this simple, low-cost marker system in routine genotyping for Bph14 in breeding populations.     

表型与分子标记相结合分析130份糖用甜菜种质的遗传多样性

旨在对甜菜种质进行比较系统的遗传多样性分析与遗传距离分析。将表型与分子标记相结合,从3 个角度分析了130 份甜菜种质的遗传多样性,并对种质进行了遗传距离的分析。130 份甜菜种质表现出比较丰富的遗传多样性,其中22 项描述性性状的多样性指数在0.0451~1.1867 之间,平均值为0.8559;23 项数值型性状的变异系数在2.94%~594.84%之间,平均值为59.10%;34 个SSR标记的多态性信息含量在0.1289~0.7847 之间,平均值为0.4887。联合使用形态标记与分子标记计算遗传距离,将130份种质分成了5个近缘组,相邻组间产糖量、块根锤度、根腐病病情指数3项性状均有显著差异。结果表明被分析的甜菜种质遗传多样性比较丰富,遗传距离分析结果有生物学意义,可为种质创新工作提供参考。     

Orientation and integration of the classical and molecular genetic maps of chromosome 11 in rice

Summary The classical genetic map and molecular map of rice chromosome 11 were oriented to facilitate the use of these maps for genetic studies and rice improvement. Three morphological markers (d-27, z-2, and la) were crossed to a rice breeding line, IRBB21, which has the Xa-21 gene for bacterial blight resistance. Three F₂ populations were analyzed with RFLP markers known to be located on chromosome 11. Segregation analysis of molecular markers and morphological markers was used to construct an RFLP map for each population. The recombination frequency between markers varied from population to population although the marker order on the maps was the same for all three populations. Based on a common set of markers mapped in the three populations, an integrated map was generated consisting of both RFLP and morphological markers. The genetic distance between markers on this map was determined by taking a weighted average of the data from the three populations. The oriented map serves as a bridge to understand the relationship between the classical and molecular linkage maps. Based on this information, the location of several genes on the classical map can be approximated with respect to RFLP markers without having to map them directly.     

Molecular fingerprinting of hybrids and assessment of genetic purity of hybrid seeds in rice using microsatellite markers

Microsatellite markers were used for fingerprinting of hybrids, assessing variation within parental lines and testing the genetic purity of hybrid seed lot in rice. Ten sequence tagged microsatellite sites (STMS) markers were employed for fingerprinting 11 rice hybrids and their parental lines. Nine STMS markers were found polymorphic across the hybrids and produced unique fingerprint for the 11 hybrids. A set of four markers (RM 206, RM 216, RM 258 and RM 263) differentiated all the hybrids from each other, which can be used as referral markers for unambiguous identification and protection of these hybrids. Cluster analysis based on Jaccard's similarity coefficient using UPGMA grouped the hybrids into three clusters. Within the cluster all the hybrids shared a common cytoplasmic male sterile line as female parent. The genetic similarity between the hybrids ranged from 0.33 to 0.92 with an average similarity index of 0.63. The analysis of plant-to-plant variation within the parental lines of the hybrid Pusa RH 10, using informative markers indicated residual heterozygosity at two marker loci. This highlights the importance of STMS markers in maintaining the genetic purity of the parental lines. The unique value of the restorer gene linked marker for testing the genetic purity of hybrid seeds is demonstrated for the first time.