DNA遗传标记技术在林业生产中的应用

林木种子及苗木鉴别技术的准确性和可靠性是提高营林造林质量的关键环节,传统的种苗鉴别技术的局限性往往使之不能有效鉴别林木种子的品种及种源。ＤＮＡ遗传标记技术则能提供稳定、准确、可靠的种及品种特异性标记,因而成为林业生产上极具潜力的品种鉴定手段,本文综述了目前国际上ＤＮＡ遗传标记技术的研究进展,并讨论了开展我国林木种菌ＤＮＡ鉴定技术研究的若干设想。     

分子标记技术及其发展

认识到生物个体之间ＤＮＡ序列差异并以此作为标记的分子标记技术的研究始于1 980年 ,之后随着分子生物学技术的发展 ,ＤＮＡ分子标记技术也得到迅速发展 ,现在ＤＮＡ标记技术已有数十种 ,它们被广泛应用于作物遗传育种、基因组作图、基因定位、植物亲缘关系鉴别、基因库构建、基因克隆等方面。本文仅就几类主要分子标记技术作一简单介绍。1 .基于分子杂交技术的分子标记技术1 .1　限制性片段长度多态性 (ｒｅｓｔｒｉｃｔｉｏｎｆｒａｇ ｍｅｎｔｌｅｎｇｔｈｐｏｌｙｍｏｒｐｈｉｓｍ ,ＲＦＬＰ)　　ＲＦＬＰ是研究最早的分子标记技术 ,1 …     

DNA分子标记技术及其在植物遗传多样性研究中的应用

本文阐述了ＤＮＡ限制性片段长度多态性、ＤＮＡ指纹图谱、单位点小卫星、单位点微卫星及随机扩增多态ＤＮＡ等主要的ＤＮＡ遗传标记技术的基本原理和方法．综述了不同ＤＮＡ标记的优缺点及其在植物遗传多样性研究中的应用前景．     

DNA分子标记技术在玉米种子纯度鉴定中的应用

从常规鉴定、生化鉴定及分子标记技术鉴定等方面,阐述了玉米种子纯度鉴定的重要性。进一步对RAPD、RFLP、SSR和AFLP等分子标记技术在种子纯度鉴定和品种真实性分析中的应用潜力及存在问题进行比较,得出SSR分子标记技术是目前种子纯度和品种真实性鉴定中最适宜的技术。     

中国林木遗传资源利用与可持续经营状况

中国经过遗传改良的重要造林树种有100多种,全国年均提供各类林木种子2300万kg,年均生产各类良种壮苗约130亿株。林木良种在生产上的应用产生了明显的综合增益,其中用材林平均生长增益达10%~30%,经济林平均产量增益达15%~68%。中国每年进口林木种子15万kg以上,涉及50多个树种;每年出口林木种子30万kg和苗木400多种。近10年来,中国林木遗传资源的可持续经营和利用已取得了明显的进步,但与一些发达国家相比还存在一定差距。今后,应优先考虑对已保存的林木遗传资源的维护和资金补贴,加强种苗市场监管和信息服务,进一步提高林木良种的基地供种率和良种使用率。     

细胞质遗传并非都是母系遗传

细胞质遗传一般表现为具母系遗传的特征。随着ＤＮＡ分子标记技术的发展和应用,人们已发现在动物及植物中均存在有低频的线粒体ＤＮＡ单亲父系遗传及双亲遗传的现象,对质体ＤＮＡ遗传的研究表明,被子植物的质体ＤＮＡ大多表现为母系遗传。而裸子植物的质体ＤＮＡ则主要表现为父系遗传的方式,同时也出现存在其它的遗传规律。     

分子标记及其在植物遗传育种中的应用

遗传标记可以说是生物群体中可识别的遗传多态性的一种表现形式。随着遗传研究特别是遗传作图的不断深入,遗传标记已从传统的以等位基因的表型识别为基础的形态标记、以染色体的结构和数目为特征的细胞学标记,及具有组织、发育及物种特异性的同工酶标记,拓展到目前已广泛应用的以ＤＮＡ多态性为基础的分子标记技术。现代分子标记技术的出现和发展为植物遗传育种研究的许多领域注入了新的活力。本文着重就目前植物遗传育种中所应用的一些主要分子标记技术及其应用作一概述。１　常用的分子标记技术自８０年代初有人提出用ＲＦＬＰ作为遗传…     

分子遗传标记技术在中药材鉴定中的应用

分子遗传标记技术在中药材品种鉴别中的应用是中药现代化研究的一个重要方面,本文回顾国内外中药分子标记鉴定研究的历史及现状,分析了目前分子标记在中药鉴定应用中存在的问题并提出建议,进而展望这项技术应用的前景。中药材质量的规范化,为中药材遗传图谱的建立指明方向。     

SSR分子标记在烟草研究中的应用进展

SSR分子标记技术作为最常用的分子标记技术之一,该标记技术重复性好,结果可靠,近年来在烟草遗传育种中展示了广阔的应用前景,是应用潜力较大的分子标记技术。介绍了SSR分子标记的原理及其分布特征,对其在烟草基因定位及分子标记辅助选择、种质资源研究、遗传图谱构建及种子纯度及真伪鉴定研究中的应用等方面进行了综述,并探讨了SSR分子标记技术在烟草遗传育种中的应用前景,以期为烟草SSR分子标记技术的研究提供参考。     

关于植物随机引物扩增多态性DNA标记的可靠性问题

随机扩增多态性ＤＮＡ（ｒａｎｄｏｍａｍｐｌｉｆｉｅｄｐｏｌｙｍｏｒｐｈｉｃＤＮＡ,ＲＡＰＤ）技术是由Ｗｉｌｌｉａｍｓ等［１］首先创立的一种ＤＮＡ分子标记技术。由于ＲＡＰＤ技术具有快速、简便、通用性好,对ＤＮＡ的需求量小,质量要求低等优点,一经问世,就受到了广泛的注意,并被成功地用于动物、植物、人和微生物的遗传多样性检测、基因定位、品系鉴定、医学诊断、遗传图谱构建和系统学研究等。在ＲＡＰＤ分子标记的应用研究过程中,人们得出了两种截然不同的结论：一部分人认为ＲＡＰＤ标记的遗传符合孟德尔分离规律,稳定…     

Application of AFLP™-based molecular markers to breeding of Populus spp.

Molecular marker technologies have eased and potentiated genetic analysis of plants and have become an extremely useful tool in forest tree breeding. The information provided by molecular markers has made it possible to acquire further knowledge about the structure and organization of plant genomes as well as about the evolution of these plant genomes through phylogenetic analysis. Using Populus spp. as a model tree, this paper aims at showing and discussing the possible applications of AFLP, a high-density DNA marker technology developed by Keygene N.V. (Wageningen, The Netherlands). Applications include: (i) AFLP analysis of the disease resistance against Melampsora larici-populina using bulked-segregant analysis, (ii) AFLP fingerprinting for identification and taxonomic analysis of individual trees, and (iii) AFLP-based mapping strategies in Populus.Abbreviations AFLP amplified fragment length polymorphism - RFLP restriction fragment length polymorphism - PCR polymerase chain reaction - QTL quantitative trait loci - RAPD random amplified polymorphic DNA     

Forest biotechnology: Innovative methods, emerging opportunities

Summary The productivity of plantation forests is essential to meet the future world demand for wood and wood products in a sustainable fashion and in a manner that preserves natural stands and biodiversity. Plantation forestry has enormously benefited from development and implementation of improved silvicultural and forest management practices during the past century. A second wave of improvements has been brought about by the introduction of new germplasm developed through genetics and breeding efforts for both hardwood and conifer tree species. Coupled with the genetic gains achieved through tree breeding, the emergence of new biotechnological approaches that span the fields of plant developmental biology, genetic transformation, and discovery of genes associated with complex multigenic traits have added a new dimension to forest tree improvement programs. Significant progress has been made during the past five years in the area of plant regeneration via organogenesis and somatic embryogenesis (SE) for economically important tree species. These advances have not only helped the development of efficient gene transfer techniques, but also have opened up avenues for deployment of new high-performance clonally replicated planting stocks in forest plantations. One of the greatest challenges today is the ability to extend this technology to the most elite germplasm, such that it becomes an, economically feasible means for large-scale production and delivery of improved planting stock. Another challenge will be the ability of the forestry research community to capitalize rapidly on current and future genomics-based elucidation of the underlying mechanisms for important but complex phenotypes. Advancements in gene cloning and genomics technology in forest trees have enabled the discovery and introduction of value-added traits for wood quality and resistance to biotic and abiotic stresses into improved genotypes. With these technical advancements, it will be necessary for reliable regulatory infrastructures and processes to be in place worldwide for testing and release of trees improved through biotechnology. Commercialization of planting stocks, as new varieties generated through clonal propagation and advanced breeding programs or as transgenic trees with high-value traits, is expected in the near future, and these trees will enhance the quality and productivity of our plantation forests.     

Applications of DNA genetic markers to the study of plant growth and development

DNA genetic markers, such as restriction fragment length polymorphisms (RFLPs) and random amplified polymorphic DNA markers (RAPDs), are powerful tools for studying the genetics of plant growth and development. DNA markers are defined sequences of DNA that can be used in traditional linkage mapping. Using DNA marker technology, scientists can uncover relationships between cloned cDNA sequences and classically characterized genes. DNA markers make it possible to dissect the contributions of multiple genetic loci underlying complex developmental processes. Moreover, changes in genome organization that occur during development or in response to environmental signals can be monitored using RFLP technology. In the future, it may be possible to clone any gene based solely on its map position. This will involve the use of tightly linked DNA markers as entry points for chromosome walking, in which a series of overlapping genomic clones reaching from the tightly linked DNA marker to the gene of interest are identified.     

Conversion of AFLP markers to high-throughput markers in a complex polyploid,sugarcane

The application of DNA markers linked to traits of commercial value in sugarcane may increase the efficiency of sugarcane breeding. The majority of markers generated for quantitative trait locus mapping in sugarcane have been single sequence repeats or AFLPs (amplified fragment length polymorphisms). Since AFLP markers are not adapted for large-scale implementation in plant breeding, our objective was to assess the feasibility of converting AFLP markers to fast, cheap and reliable PCR-based assays in a complex polyploid, sugarcane. Three AFLP markers were selected on the basis of an association to resistance to the fungal pathogen Ustilago scitaminea, the causal agent of smut in sugarcane. We developed an approach which enabled the identification of polymorphisms in these AFLP markers. Towards this goal, we employed GenomeWalking and 454 sequencing to isolate sequences adjacent to the linked AFLP markers and identify SNP (single nucleotide polymorphisms) haplotypes present in the homo(eo)logous chromosomes of sugarcane. One AFLP marker was converted to a cleavage amplified polymorphic sequence marker, another to a SCAR (sequence characteristered amplified region) marker and the final AFLP marker to a SNP PCR-based assay. However, validation of each of the markers in 240 genotypes resulted in 99, 90 and 60% correspondence with the original AFLP marker. These experiments indicate that even in a complex polyploid such as sugarcane, polymorphisms identified by AFLP can be converted to high-throughput marker systems, but due to the complexity this would only be carried out for high-value markers. In some cases, the polymorphisms identified are not transferable to more sequence-specific PCR applications.     

Genotyping single nucleotide polymorphisms in barley by tetra-primer ARMS-PCR.

Single nucleotide polymorphisms (SNPs) are the most abundant form of DNA polymorphism. These polymorphisms can be used in plants as simple genetic markers for many breeding applications, for population studies, and for germplasm fingerprinting. The great increase in the available DNA sequences in the databases has made it possible to identify SNPs by "database mining", and the single most important factor preventing their widespread use appears to be the genotyping cost. Many genotyping platforms rely on the use of sophisticated, automated equipment coupled to costly chemistry and detection systems. A simple and economical method involving a single PCR is reported here for barley SNP genotyping. Using the tetra-primer ARMS-PCR procedure, we have been able to assay unambiguously five SNPs in a set of 132 varieties of cultivated barley. The results show the reliability of this technique and its potential for use in low- to moderate-throughput situations; the association of agronomically important traits is discussed.     

Inferring species trees directly from biallelic genetic markers: bypassing gene trees in a full coalescent analysis

The multispecies coalescent provides an elegant theoretical framework for estimating species trees and species demographics from genetic markers. However, practical applications of the multispecies coalescent model are limited by the need to integrate or sample over all gene trees possible for each genetic marker. Here we describe a polynomial-time algorithm that computes the likelihood of a species tree directly from the markers under a finite-sites model of mutation effectively integrating over all possible gene trees. The method applies to independent (unlinked) biallelic markers such as well-spaced single nucleotide polymorphisms, and we have implemented it in SNAPP, a Markov chain Monte Carlo sampler for inferring species trees, divergence dates, and population sizes. We report results from simulation experiments and from an analysis of 1997 amplified fragment length polymorphism loci in 69 individuals sampled from six species of Ourisia (New Zealand native foxglove).     

Transcript-specific, single-nucleotide polymorphism discovery and linkage analysis in hexaploid bread wheat (Triticum aestivum L.)

Food security is a global concern and substantial yield increases in cereal crops are required to feed the growing world population. Wheat is one of the three most important crops for human and livestock feed. However, the complexity of the genome coupled with a decline in genetic diversity within modern elite cultivars has hindered the application of marker‐assisted selection (MAS) in breeding programmes. A crucial step in the successful application of MAS in breeding programmes is the development of cheap and easy to use molecular markers, such as single‐nucleotide polymorphisms. To mine selected elite wheat germplasm for intervarietal single‐nucleotide polymorphisms, we have used expressed sequence tags derived from public sequencing programmes and next‐generation sequencing of normalized wheat complementary DNA libraries, in combination with a novel sequence alignment and assembly approach. Here, we describe the development and validation of a panel of 1114 single‐nucleotide polymorphisms in hexaploid bread wheat using competitive allele‐specific polymerase chain reaction genotyping technology. We report the genotyping results of these markers on 23 wheat varieties, selected to represent a broad cross‐section of wheat germplasm including a number of elite UK varieties. Finally, we show that, using relatively simple technology, it is possible to rapidly generate a linkage map containing several hundred single‐nucleotide polymorphism markers in the doubled haploid mapping population of Avalon × Cadenza.     

Sequence-related amplified polymorphism (SRAP) molecular marker system and its applications in crop improvement

Plant biotechnology has great potential for improving target traits in crops. This can be achieved by the production of transgenic crops and marker-assisted breeding. Marker-assisted breeding has gained momentum in recent times since it does not need biosafety regulations. Several kind of molecular markers are available for use in crop breeding, such as restriction fragment length polymorphism, microsatellites, sequence characterized amplified region, sequence-tagged site, inter-simple sequence repeat amplification, amplified fragment length polymorphism and single nucleotide polymorphism. Sequence-related amplified polymorphism is a novel molecular marker system which is based on open reading frames (ORFs) developed from genome sequence data of Arabidopsis. It provides a unique combination of forward and reverse primers which can be selected arbitrarily giving a large number of primer combinations. Since this is an ORF-based marker system, it targets functional genes and has potential for their application in crop breeding. This marker system was first used and demonstrated by Li and Quiros in Brassica oleracea in 2001, and since then there have been several reports in different plant species ranging from field crops to tree species for assessing genetic diversity, mapping and tagging of genes, hybrid identification and sex determination. This review provides an overview of SRAP markers and their applications in crop improvement.     

西瓜核心种质的AFLP指纹图谱和SCAR标记

西瓜(Citrullus lanatus (Thunb.) Mansf.)种质资源的鉴定与评价是对其有效利用的基础.以往的研究表明, 西瓜是一种遗传资源特别狭窄的作物,在用同工酶、RAPD及SSR技术对西瓜种质资源进行鉴定时,发现很难将品种完全区分开来.本研究利用高效可靠的AFLP技术,对30个西瓜核心种质材料进行了遗传分析,最终建立了这30个材料的DNA指纹图谱.在该图谱中,每个材料均有其独特的"指纹",材料之间可以相互区分开来.为了进一步利用AFLP分子标记,将重要抗病种质材料"PI296341"的AFLP特异带转化成了生产上可以直接利用的SCAR标记.     

Reproducibility testing of RAPD,AFLP and SSR markers in plants by a network of European laboratories

A number of PCR-based techniques can be used to detect polymorphisms in plants. For their wide-scale usage in germplasm characterisation and breeding it is important that these marker technologies can be exchanged between laboratories, which in turn requires that they can be standardised to yield reproducible results, so that direct collation and comparison of the data are possible. This article describes a network experiment involving several European laboratories, in which the reproducibility of three popular molecular marker techniques was examined: random-amplified fragment length polymorphism (RAPD), amplified fragment length polymorphism (AFLP) and sequence-tagged microsatellites (SSR). For each technique, an optimal system was chosen, which had been standardised and routinely used by one laboratory. This system (genetic screening package) was distributed to different participating laboratories in the network and the results obtained compared with those of the original sender. Different experiences were gained in this exchange experiment with the different techniques. RAPDs proved difficult to reproduce. For AFLPs, a single-band difference was observed in one track, whilst SSR alleles were amplified by all laboratories, but small differences in their sizing were obtained.