Choosing and using a plant DNA barcode

The main aim of DNA barcoding is to establish a shared community resource of DNA sequences that can be used for organismal identification and taxonomic clarification. This approach was successfully pioneered in animals using a portion of the cytochrome oxidase 1 (CO1) mitochondrial gene. In plants, establishing a standardized DNA barcoding system has been more challenging. In this paper, we review the process of selecting and refining a plant barcode; evaluate the factors which influence the discriminatory power of the approach; describe some early applications of plant barcoding and summarise major emerging projects; and outline tool development that will be necessary for plant DNA barcoding to advance.     

藻类DNA条形码研究进展

DNA barcode,又称为DNA条形码,是指利用短的标准DNA序列的核苷酸多样性进行物种的鉴定和快速识别.目前该方法在动物分类研究中应用广泛,其中线粒体的细胞色素c氧化酶亚基1(cytochrome c oxidase subunit 1,COI或cox 1)基因中的约700bp长度的一段被用来作为标准DNA片段.在陆地植物条形码研究中,生命-植物条形码联盟会(Consortium for the Barcode of Life-Plant Working Group,CBOL-Plant Working Group)近期推荐将植物叶绿体中的两个基因片段rbcL+ matK作为初步的陆生植物条形码,此组合能在70％的程度上进行植物物种的鉴别.在海藻的分类研究中,DNA条形码的应用较少,已有的研究主要集中在硅藻、红藻和褐藻,尚没有学者明确提出适合藻类的DNA条形码.总结了能够作为藻类DNA条形码的序列特点、应用流程及分析方法,综述了DNA条形码在藻类中的研究现状和存在的问题,展望了藻类DNA条形码的应用前景.     

DNA条形码在鞘翅目昆虫分子系统学研究中的应用

近年来,DNA条形码(DNA Barcoding)技术已经成为生物分类学研究中备受关注的新型技术,并在鞘翅目昆虫系统发育研究中得到广泛应用。本文总结了鞘翅目昆虫DNA条形码研究所用COⅠ基因序列,概述了DNA条形码在鞘翅目昆虫的物种分类鉴定、发现新种和隐存种、系统发育关系研究等方面的应用,并对DNA条形码研究技术新进展和标准序列筛选需要注意的问题进行了讨论。     

蒟蒻薯属 (薯蓣科) 植物DNA条形码研究

蒟蒻薯属（Tacca）植物种间在形态上差别不大,导致分类上存在一定的困难。DNA条形码是一种利用短的DNA标准片段来鉴别和发现物种的方法。本研究利用核基因ITS片段和叶绿体基因trnH psbA, rbcL, matK片段对蒟蒻薯属6个种的DNA条形码进行研究,对4个DNA片段可用性,种内种间变异,barcode gap进行了分析,采用Tree based和BBA两种方法比较不同序列的鉴定能力。结果显示：单片段ITS正确鉴定率最高,片段组合rbcL+matK正确鉴定率最高。支持CBOL植物工作组推荐的条码组合rbcL+matK可作为蒟蒻薯属物种鉴定的标准条码,建议ITS片段作为候选条码。丝须蒟蒻薯Tacca integrifolia采自西藏的居群与马来西亚居群形成了2个不同的遗传分支,且两者在形态上也存在一定的差异,很可能是一个新种。     

姜科砂仁属植物DNA条形码序列的筛选

砂仁属（Amomum）隶属于姜科,全属约150种,我国有39种。该属多种植物可作药物或香料,但目前砂仁属的分类还不清楚,准确鉴定物种有很大难度。本研究利用DNA barcoding技术,对砂仁属50种121个个体的matK、rbcL-a、trnH-psbA序列及其不同组合进行比较,用Taxon DNA计算种间、种内bar-coding gap,运用相似法的BLASTn计算条码的正确鉴定率,筛选适合砂仁属的条码片段。结果显示：所有条码的barcoding gap均不存在;matK的正确鉴定率高于trnH-psbA和rbcL-a,联合片段的条码正确鉴定率高于单片段条码,三个片段联合条码的正确鉴定率最高。因此,推荐matK＋rbcL-a＋trnH-psbA作为砂仁属物种鉴定的候选条码。     

Applying plant DNA barcodes to identify species of Parnassia (Parnassiaceae)

DNA barcoding is a technique to identify species by using standardized DNA sequences. In this study, a total of 105 samples, representing 30 Parnassia species, were collected to test the effectiveness of four proposed DNA barcodes (rbcL, matK, trnH-psbA and ITS) for species identification. Our results demonstrated that all four candidate DNA markers have a maximum level of primer universality and sequencing success. As a single DNA marker, the ITS region provided the highest species resolution with 86.7%, followed by trnH-psbA with 73.3%. The combination of the core barcode regions, matK+rbcL, gave the lowest species identification success (63.3%) among any combination of multiple markers and was found unsuitable as DNA barcode for Parnassia. The combination of ITS+trnH-psbA achieved the highest species discrimination with 90.0% resolution (27 of 30 sampled species), equal to the four-marker combination and higher than any two or three marker combination including rbcL or matK. Therefore, matK and rbcL should not be used as DNA barcodes for the species identification of Parnassia. Based on the overall performance, the combination of ITS+trnH-psbA is proposed as the most suitable DNA barcode for identifying Parnassia species. DNA barcoding is a useful technique and provides a reliable and effective mean for the discrimination of Parnassia species, and in combination with morphology-based taxonomy, will be a robust approach for tackling taxonomically complex groups. In the light of our findings, we found among the three species not identified a possible cryptic speciation event in Parnassia.     

Comprehensive DNA barcode coverage of North American birds

DNA barcoding seeks to assemble a standardized reference library for DNA-based identification of eukaryotic species. The utility and limitations of this approach need to be tested on well-characterized taxonomic assemblages. Here we provide a comprehensive DNA barcode analysis for North American birds including 643 species representing 93% of the breeding and pelagic avifauna of the USA and Canada. Most (94%) species possess distinct barcode clusters, with average neighbour-joining bootstrap support of 98%. In the remaining 6%, barcode clusters correspond to small sets of closely related species, most of which hybridize regularly. Fifteen (2%) currently recognized species are comprised of two distinct barcode clusters, many of which may represent cryptic species. Intraspecific variation is weakly related to census population size and species age. This study confirms that DNA barcoding can be effectively applied across the geographical and taxonomic expanse of North American birds. The consistent finding of constrained intraspecific mitochondrial variation in this large assemblage of species supports the emerging view that selective sweeps limit mitochondrial diversity.     

Testing four proposed barcoding markers for the identification of species within Ligustrum L.(Oleaceae)

DNA barcoding is a biological technique that uses short and standardized genes or DNA regions to facilitate species identification. DNA barcoding has been used successfully in several animal and plant groups. Ligustrum (Oleaceae) species occur widely throughout the world and are used as medicinal plants in China. Therefore, the accurate identification of species in this genus is necessary. Four potential DNA barcodes, namely the nuclear ribosomal internal transcribed spacer (ITS) and three chloroplast (cp) DNA regions (rbcL, marK, and trnH-psbA),were used to differentiate species within Ligustrum. BLAST, character-based method, tree-based methods and TAXONDNA analysis were used to investigate the molecular identification capabilities of the chosen markers for discriminating 92 samples representing 20 species of this genus. The results showed that the ITS sequences have the most variable information, followed by trnH-psbA, matK, and rbcL. All sequences of the four regions correctly identified the species at the genus level using BLAST alignment. At the species level, the discriminating power of rbcL, matK, trnH-psbA and ITS based on neighbor-joining (NJ) trees was 36.8%, 38.9%, 77.8%, and 80%,respectively. Using character-based and maximum parsimony (MP) tree methods together, the discriminating ability of trnH-psbA increased to 88.9%. All species could be differentiated using ITS when combining the NJ tree method with character-based or MP tree methods. Overall, the results indicate that DNA barcoding is an effective molecular identification method for Ligustrum species. We propose the nuclear ribosomal ITS as a plant barcode for plant identification and trnH-psbA as a candidate barcode sequence.     

Testing four proposed barcoding markers for the identification of species within Ligustrum L. (Oleaceae)

DNA barcoding is a biological technique that uses short and standardized genes or DNA regions to facilitate species identification. DNA barcoding has been used successfully in several animal and plant groups. Ligustrum (Oleaceae) species occur widely throughout the world and are used as medicinal plants in China. Therefore, the accurate identification of species in this genus is necessary. Four potential DNA barcodes, namely the nuclear ribosomal internal transcribed spacer (ITS) and three chloroplast (cp) DNA regions (rbcL, matK, and trnH–psbA), were used to differentiate species within Ligustrum. BLAST, character-based method, tree-based methods and TAXONDNA analysis were used to investigate the molecular identification capabilities of the chosen markers for discriminating 92 samples representing 20 species of this genus. The results showed that the ITS sequences have the most variable information, followed by trnH–psbA, matK, and rbcL. All sequences of the four regions correctly identified the species at the genus level using BLAST alignment. At the species level, the discriminating power of rbcL, matK, trnH–psbA, and ITS based on neighbor-joining (NJ) trees was 36.8%, 38.9%, 77.8%, and 80%, respectively. Using character-based and maximum parsimony (MP) tree methods together, the discriminating ability of trnH–psbA increased to 88.9%. All species could be differentiated using ITS when combining the NJ tree method with character-based or MP tree methods. Overall, the results indicate that DNA barcoding is an effective molecular identification method for Ligustrum species. We propose the nuclear ribosomal ITS as a plant barcode for plant identification and trnH–psbA as a candidate barcode sequence.     

DNA barcoding of Pedicularis L.(Orobanchaceae): Evaluating four universal barcode loci in a large and hemiparasitic genus

One application ofDNA barcoding is species identification based on sequences of a short and standardized DNA region.In plants,various DNA regions,alone or in combination,have been proposed and investigated,but consensus on a universal plant barcode remains elusive.In this study,we tested the utility of four candidate barcoding regions (rbcL,matK,trnH-psbA,and internal transcribed spacer (ITS)) as DNA barcodes for discriminating species in a large and hemiparasitic genus Pedicularis (Orobanchaceae).Amplification and sequencing was successful using single primer pairs for rbcL,trnH-psbA,and ITS,whereas two primer pairs were required for matK.Patterns of sequence divergence commonly showed a “barcoding gap”,that is,a bimodal frequency distribution of pairwise distances representing genetic diversity within and between species,respectively Considering primer universality,ease of amplification and sequencing,and performance in discriminating species,we found the most effective single-region barcode for Pedicularis to be ITS,and the most effective two-region barcode to be rbcL +ITS.Both discriminated at least 78％ of the 88 species and correctly identified at least 89％ of the sequences in our sample,and were effective in placing unidentified samples in known species groups.Our results suggest that DNA barcoding has the potential to aid taxonomic research in Pedicularis,a species-rich cosmopolitan clade much in need of revision,as well as ecological studies in its center of diversity,the Hengduan Mountains region of China.     

Molecular-based rapid inventories of sympatric diversity: A comparison of DNA barcode clustering methods applied to geography-based vs clade-based sampling of amphibians

Molecular markers offer a universal source of data for quantifying biodiversity. DNA barcoding uses a standardized genetic marker and a curated reference database to identify known species and to reveal cryptic diversity within well-sampled clades. Rapid biological inventories, e.g. rapid assessment programs (RAPs), unlike most barcoding campaigns, are focused on particular geographic localities rather than on clades. Because of the potentially sparse phylogenetic sampling, the addition of DNA barcoding to RAPs may present a greater challenge for the identification of named species or for revealing cryptic diversity. In this article we evaluate the use of DNA barcoding for quantifying lineage diversity within a single sampling site as compared to clade-based sampling, and present examples from amphibians. We compared algorithms for identifying DNA barcode clusters (e.g. species, cryptic species or Evolutionary Significant Units) using previously published DNA barcode data obtained from geography-based sampling at a site in Central Panama, and from clade-based sampling in Madagascar. We found that clustering algorithms based on genetic distance performed similarly on sympatric as well as clade-based barcode data, while a promising coalescent-based method performed poorly on sympatric data. The various clustering algorithms were also compared in terms of speed and software implementation. Although each method has its shortcomings in certain contexts, we recommend the use of the ABGD method, which not only performs fairly well under either sampling method, but does so in a few seconds and with a user-friendly Web interface.     

DNA barcoding to map the microbial communities: current advances and future directions

During the last two decades, the DNA barcode development towards microbial community has increased dramatically. DNA barcode development is related to error-free and quick species identification which aid in understanding the microbial biodiversity, as well as the diseases related to microbial species. Here, we seek to evaluate the so-called barcoding initiatives for the microbial communities and the emerging trends in this field. In this paper, we describe the development of DNA marker-based DNA barcoding system, comparison between routine species identification and DNA barcode, and microbial biodiversity and DNA barcode for microbial communities. Two major topics, such as the molecular diversity of viruses and barcode for viruses have been discussed at the same time. We demonstrate the current status and the maker of DNA barcode for bacteria, algae, fungi, and protozoa. Furthermore, we argue about the promises, limitations, and present and future challenges of microbial barcode development.     

DNA条形码：物种分类和鉴定技术

当前,一项称为“生命的条形码”计划正在欧美等国展开,其目的是实现对地球上现存的约1000万物种进行快速和准确的鉴定。DNA条形码是一种利用短的DNA序列对物种进行鉴定的技术。对DNA条形码的概念和原理进行了介绍,举例说明了其在物种分类、遗传多样性及物种鉴定研究中广泛的利用价值,阐述了当前该领域的研究现状,对未来的发展方向进行了展望。     

真菌DNA条形码技术研究进展

DNA条形码(DNA barcoding)技术作为一门新兴的物种鉴定方法以其灵敏、精确、方便和客观的优势,在动植物和微生物的分类鉴定中已经得到广泛应用.真菌鉴定中常用作标准条形码的是核核糖体DNA内转录间隔区(Internal transcribed spacer,ITS),如今也有一些新型条形码被发现和应用到实际操作中,如微条形码、ND6、EF3.本文对DNA条形码技术的产生和发展做出了总结,通过研究其在真菌中应用的实际案例分析了DNA条形码技术的优缺点及发展趋势,并指出DNA条形码技术将以全新的视角来弥补传统分类学的不足,最终实现生物自身的序列变异信息与现有形态分类学的结合.     

DNA条形码分析方法研究进展

DNA条形码是一段短的、标准化的DNA序列,DNA条形码技术通过对DNA条形码序列分析实现物种的有效鉴定.随着生物DNA条形码序列的大量测定,DNA条形码分析方法得到迅速发展,推动了其在生物分子鉴定中的应用.2003年以来,DNA条形码技术已广泛应用于动物、植物和真菌等物种的鉴定,并有力地推动了生物分类学、生物多样性和生态学等学科的发展.本文在综述DNA条形码技术的基础上,总结了5类主要的DNA条形码分析方法,即基于遗传距离的分析、基于遗传相似度的分析、基于系统发育树的分析、基于序列特征的分析和基于统计分类法的分析,并进一步展望了DNA条形码技术的发展与应用.     

悬钩子属DNA条形码通用序列的初步筛选

为了建立悬钩子属（Rubus）植物的DNA条形码分子鉴定技术,筛选获得适用于悬钩子属植物的通用条形码序列。该研究基于GenBank数据对ITS、ITS2、matK、rbcL、trnH-psbA、trnL-trnF 6个DNA条形码序列进行了遗传变异、barcoding gap、建树等评估分析。结果显示,trnH-psbA、matK、rbcL、rtnL-trnF的种内变异与种间变异差异较大,变异分辨率分别为97.32%、83.33%、79.07%、64.95%,存在较大的barcoding gap;NJ一致树分析显示,matK的单系性比例最高（67%）,其次为trnH-psbA（64%）,rtnL-trnF（43%）,rbcL（30%）。结果表明,悬钩子属植物的matK与trnH-psbA序列种内变异与种间变异差异较大,能较好地区分不同物种,具有较大的鉴定潜力。建议将matK和trnH-psbA作为悬钩子属植物鉴定的核心条形码序列,rtnL-trnF、rbcL作为辅助条形码序列。     

植物DNA条形码与生物多样性数据共享平台构建

DNA条形码基于较短的DNA序列实现物种的快速、准确鉴定, 不仅加快了全球生物物种的鉴定和分类步伐, 也为生物多样性的管理、保护和可持续利用提供了新思路和研究方法。植物DNA条形码标准数据库的不断完善, 将使植物多样性信息的快速获取成为可能; 将不同类型数据资源整合、共享和利用, 构建植物DNA条形码数据共享平台, 是满足公众对物种准确鉴定和快速认知的重要支撑。本文介绍了近年来植物DNA条形码的研究进展; 植物DNA条形码参考数据库的研发现状和存在的问题。结合上述问题, 围绕“大数据”时代背景, 对如何管理和使用好海量的植物信息, 如何构建数据共享平台提出了一些设想: (1)数据共享平台的元数据应尽可能翔实、丰富、准确和多关联; (2)数据标准应统一规范; (3)查询入口方便、迅速、多样, 易于管理, 便于实现更大程度的数据共享和全球化的合作交流。     

DNA barcoding of Scandinavian birds reveals divergent lineages in trans-Atlantic species

Birds are a taxonomically well-described group of animals, yet DNA barcoding, i.e., the molecular characterization of species using a standardized genetic marker, has revealed unexpected patterns of genetic divergences among North American birds. We performed a comprehensive COI (cytochrome c oxidase subunit I) barcode survey of 296 species of Scandinavian birds, and compared genetic divergences among 78 trans-Atlantic species whose breeding ranges include both Scandinavia and North America. Ninety-four percent of the Scandinavian species showed unique barcode clusters; the remaining 6% had overlapping barcodes with one or more congeneric species, which may reflect incomplete lineage sorting or a single gene pool. Four species showed large intra-specific divergences within Scandinavia, despite no apparent morphological differentiation or indications of reproductive isolation. These cases may reflect admixture of previously isolated lineages, and may thus warrant more comprehensive phylogeographic analyses. Nineteen (24%) of 78 trans-Atlantic species exhibited divergent genetic clusters which correspond with regional subspecies. Three of these trans-Atlantic divergences were paraphyletic. Our study demonstrates the effectiveness of COI barcodes for identifying Scandinavian birds and highlights taxa for taxonomic review. The standardized DNA barcoding approach amplified the power of our regional studies by enabling independently obtained datasets to be merged with the established avian barcode library.     

线粒体COⅠ基因在昆虫DNA条形码中的研究与应用

自2003年DNA条形码(DNA barcodes)概念出现以来,DNA条形码技术(DNA barcoding)受到生物分类学领域普遍关注,线粒体细胞色素氧化酶亚基I(mtDNACOⅠ)被用作动物类群的主要条形码序列,基于该基因片段的昆虫条形码研究在国内外广泛开展。本文在概述DNA条形码、条形码技术及已开展的昆虫条形码研究计划的基础上,总结了昆虫mtDNACOⅠ条形码及其技术在发现和描述隐种、种类分子鉴定以及系统发育等方面的研究进展,分析了细胞核线粒体假基因(Numts)对mtDNACOⅠ条形码扩增的影响,提出检测和避免Numts的方法,并对DNA条形码技术的进一步研究和应用进行了讨论和展望。     

From barcodes to genomes: extending the concept of DNA barcoding

DNA barcoding has had a major impact on biodiversity science. The elegant simplicity of establishing massive scale databases for a few barcode loci is continuing to change our understanding of species diversity patterns, and continues to enhance human abilities to distinguish among species. Capitalizing on the developments of next generation sequencing technologies and decreasing costs of genome sequencing, there is now the opportunity for the DNA barcoding concept to be extended to new kinds of genomic data. We illustrate the benefits and capacity to do this, and also note the constraints and barriers to overcome before it is truly scalable. We advocate a twin track approach: (i) continuation and acceleration of global efforts to build the DNA barcode reference library of life on earth using standard DNA barcodes and (ii) active development and application of extended DNA barcodes using genome skimming to augment the standard barcoding approach.