Googling DNA sequences on the World Wide Web

Background: New web-based technologies provide an excellent opportunity for sharing and accessing information and using web as a platform for interaction and collaboration. Although several specialized tools are available for analyzing DNA sequence information, conventional webbased tools have not been utilized for bioinformatics applications. We have developed a novel algorithm and implemented it for searching species-specific genomic sequences, DNA barcodes, by using popular web-based methods such as Google. Results: We developed an alignment independent character based algorithm based on dividing a sequence library (DNA barcodes) and query sequence to words. The actual search is conducted by conventional search tools such as freely available Google Desktop Search. We implemented our algorithm in two exemplar packages. We developed pre and post-processing software to provide customized input and output services, respectively. Our analysis of all publicly available DNA barcode sequences shows a high accuracy as well as rapid results. Conclusion: Our method makes use of conventional web-based technologies for specialized genetic data. It provides a robust and efficient solution for sequence search on the web. The integration of our search method for large-scale sequence libraries such as DNA barcodes provides an excellent web-based tool for accessing this information and linking it to other available categories of information on the web

Design and applicability of DNA arrays and DNA barcodes in biodiversity monitoring

<p>Abstract</p> <p>Background</p> <p>The rapid and accurate identification of species is a critical component of large-scale biodiversity monitoring programs. DNA arrays (micro and macro) and DNA barcodes are two molecular approaches that have recently garnered much attention. Here, we compare these two platforms for identification of an important group, the mammals.</p> <p>Results</p> <p>Our analyses, based on the two commonly used mitochondrial genes cytochrome <it>c </it>oxidase I (the standard DNA barcode for animal species) and cytochrome b (a common species-level marker), suggest that both arrays and barcodes are capable of discriminating mammalian species with high accuracy. We used three different datasets of mammalian species, comprising different sampling strategies. For DNA arrays we designed three probes for each species to address intraspecific variation. As for DNA barcoding, our analyses show that both cytochrome <it>c </it>oxidase I and cytochrome b genes, and even smaller fragments of them (mini-barcodes) can successfully discriminate species in a wide variety of specimens.</p> <p>Conclusion</p> <p>This study showed that DNA arrays and DNA barcodes are valuable molecular methods for biodiversity monitoring programs. Both approaches were capable of discriminating among mammalian species in our test assemblages. However, because designing DNA arrays require advance knowledge of target sequences, the use of this approach could be limited in large scale monitoring programs where unknown haplotypes might be encountered. DNA barcodes, by contrast, are sequencing-based and therefore could provide more flexibility in large-scale studies.</p

Stepwise implementation of high-throughput sequencing metabarcoding to estuarine macrobenthic communities

This work was supported by FEDER through POFC-COMPETE by national funds from ‘Fundação para a Ciência e a Tecnologia (FCT)’ in the scope of the grant FCOMP-01-0124-FEDER-015429 and also by the strategic programme UID/BIA/04050/2013 (POCI-01- 0145-FEDER-007569) also funded by national funds through the FCT I.P. and by the ERDF through the COMPETE2020 - Programa Operacional Competitividade e Internacionalização (POCI). Jorge Lobo was supported by a PhD fellowship (SFRH/BD/69750/2010) from FCT.info:eu-repo/semantics/publishedVersio

DNA metabarcoding for high-throughput monitoring of estuarine macrobenthic communities

Morphology-based profling of benthic communities has been extensively applied to aquatic ecosystems’ health assessment. However, it remains a low-throughput, and sometimes ambiguous, procedure. Despite DNA metabarcoding has been applied to marine benthos, a comprehensive approach providing species-level identifcations for estuarine macrobenthos is still lacking. Here we report a combination of experimental and feld studies to assess the aptitude of COI metabarcoding to provide robust species-level identifcations for high-throughput monitoring of estuarine macrobenthos. To investigate the ability of metabarcoding to detect all species present in bulk DNA extracts, we contrived three phylogenetically diverse communities, and applied four diferent primer pairs to generate PCR products within the COI barcode region. Between 78–83% of the species in the contrived communities were recovered through HTS. Subsequently, we compared morphology and metabarcoding-based approaches to determine the species composition from four distinct estuarine sites. Our results indicate that species richness would be considerably underestimated if only morphological methods were used: globally 27 species identifed through morphology versus 61 detected by metabarcoding. Although further refnement is required to improve efciency and output of this approach, here we show the great aptitude of COI metabarcoding to provide high quality and auditable species identifcations in estuarine macrobenthos monitoring.This study has been funded by the project “Te NextSea: Next generation monitoring of coastal ecosystems in a scenario of global change” (operação NORTE-01-0145-FEDER-000032), supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). JL was supported by a PhD fellowship (SFRH/BD/69750/2010) from FCT. Tis study had the fnancial support of Fundação para a Ciência e Tecnologia (FCT), through the strategic project UID/MAR/04292/2013 granted to MARE. Te authors would like to thank Stephanie Boilard (Biodiversity Institute of Ontario) for her support in the lab work.info:eu-repo/semantics/publishedVersio

Environmental Barcoding: A Next-Generation Sequencing Approach for Biomonitoring Applications Using River Benthos

Timely and accurate biodiversity analysis poses an ongoing challenge for the success of biomonitoring programs. Morphology-based identification of bioindicator taxa is time consuming, and rarely supports species-level resolution especially for immature life stages. Much work has been done in the past decade to develop alternative approaches for biodiversity analysis using DNA sequence-based approaches such as molecular phylogenetics and DNA barcoding. On-going assembly of DNA barcode reference libraries will provide the basis for a DNA-based identification system. The use of recently introduced next-generation sequencing (NGS) approaches in biodiversity science has the potential to further extend the application of DNA information for routine biomonitoring applications to an unprecedented scale. Here we demonstrate the feasibility of using 454 massively parallel pyrosequencing for species-level analysis of freshwater benthic macroinvertebrate taxa commonly used for biomonitoring. We designed our experiments in order to directly compare morphology-based, Sanger sequencing DNA barcoding, and next-generation environmental barcoding approaches. Our results show the ability of 454 pyrosequencing of mini-barcodes to accurately identify all species with more than 1% abundance in the pooled mixture. Although the approach failed to identify 6 rare species in the mixture, the presence of sequences from 9 species that were not represented by individuals in the mixture provides evidence that DNA based analysis may yet provide a valuable approach in finding rare species in bulk environmental samples. We further demonstrate the application of the environmental barcoding approach by comparing benthic macroinvertebrates from an urban region to those obtained from a conservation area. Although considerable effort will be required to robustly optimize NGS tools to identify species from bulk environmental samples, our results indicate the potential of an environmental barcoding approach for biomonitoring programs

iBarcode.org: web-based molecular biodiversity analysis

<p>Abstract</p> <p>Background</p> <p>DNA sequences have become a primary source of information in biodiversity analysis. For example, short standardized species-specific genomic regions, DNA barcodes, are being used as a global standard for species identification and biodiversity studies. Most DNA barcodes are being generated by laboratories that have an expertise in DNA sequencing but not in bioinformatics data analysis. Therefore, we have developed a web-based suite of tools to help the DNA barcode researchers analyze their vast datasets.</p> <p>Results</p> <p>Our web-based tools, available at <url>http://www.ibarcode.org</url>, allow the user to manage their barcode datasets, cull out non-unique sequences, identify haplotypes within a species, and examine the within- to between-species divergences. In addition, we provide a number of phylogenetics tools that will allow the user to manipulate phylogenetic trees generated by other popular programs.</p> <p>Conclusion</p> <p>The use of a web-based portal for barcode analysis is convenient, especially since the WWW is inherently platform-neutral. Indeed, we have even taken care to ensure that our website is usable from handheld devices such as PDAs and smartphones. Although the current set of tools available at iBarcode.org were developed to meet our own analytic needs, we hope that feedback from users will spark the development of future tools. We also welcome user-built modules that can be incorporated into the iBarcode framework.</p

Benchmarking DNA barcodes: an assessment using available primate sequences

DNA barcoding has been recently promoted as a method for both assigning specimens to known species and for discovering new and cryptic species. Here we test both the potential and the limitations of DNA barcodes by analysing a group of well-studied organisms—the primates. Our results show that DNA barcodes provide enough information to efficiently identify and delineate primate species, but that they cannot reliably uncover many of the deeper phylogenetic relationships. Our conclusion is that these short DNA sequences do not contain enough information to build reliable molecular phylogenies or define new species, but that they can provide efficient sequence tags for assigning unknown specimens to known species. As such, DNA barcoding provides enormous potential for use in global biodiversity studies

A universal DNA mini-barcode for biodiversity analysis

<p>Abstract</p> <p>Background</p> <p>The goal of DNA barcoding is to develop a species-specific sequence library for all eukaryotes. A 650 bp fragment of the cytochrome <it>c </it>oxidase 1 (CO1) gene has been used successfully for species-level identification in several animal groups. It may be difficult in practice, however, to retrieve a 650 bp fragment from archival specimens, (because of DNA degradation) or from environmental samples (where universal primers are needed).</p> <p>Results</p> <p>We used a bioinformatics analysis using all CO1 barcode sequences from GenBank and calculated the probability of having species-specific barcodes for varied size fragments. This analysis established the potential of much smaller fragments, mini-barcodes, for identifying unknown specimens. We then developed a universal primer set for the amplification of mini-barcodes. We further successfully tested the utility of this primer set on a comprehensive set of taxa from all major eukaryotic groups as well as archival specimens.</p> <p>Conclusion</p> <p>In this study we address the important issue of minimum amount of sequence information required for identifying species in DNA barcoding. We establish a novel approach based on a much shorter barcode sequence and demonstrate its effectiveness in archival specimens. This approach will significantly broaden the application of DNA barcoding in biodiversity studies.</p

Small-scale spatial variation of meiofaunal communities in Lima estuary (NW Portugal) assessed through metabarcoding

Meiofaunal organisms play a key role in estuarine ecosystems, being responsible for significant ecological processes. However, meiofauna constitutes a particularly difficult community to be monitored through conventional morphology-based approaches. New emerging tools, such as DNA metabarcoding, facilitate the access to these communities and provide an opportunity to develop routine monitoring programs. In the present study, the small-scale spatial variation of meiofaunal communities in the Lima estuary (NW Portugal) was investigated using DNA metabarcoding. The first stage of the study aimed to establish the amount of sediment sample to be used for DNA extraction and to test six primer pairs, three of them amplifying fragments from the mitochondrial cytochrome c oxidase gene (COI) and three other the nuclear ribosomal 18S rRNA gene (18S). In a subsequent stage, sediment samples were collected in four stations along an estuarine gradient (salinity ranging between 9 and 28), in which six sampling points about 4–5 m apart were considered: three in the high intertidal and three in the mid intertidal. After the DNA extraction from sediments, COI and 18S amplicon libraries were produced and sequenced in an Illumina MiSeq platform. OTUs (operational taxonomic units) recovered by either COI or 18S displayed generally high turnover in occurrence among sampling points within a station and tidal horizon, among tidal horizons within a station, and among stations of distinct salinity (approx. 60–93%). Both markers recorded little variation among stations in OTU richness and in the taxonomic composition of the most dominant groups. However, the meiofauna detected differed qualitatively between the two markers used; Amoebozoa and Cnidaria were mostly detected with COI while Ciliophora and Platyhelminthes with 18S. In addition, the structure of the meiofauna community diverged significantly among stations and was strongly influenced by salinity and sediment features. Globally, results indicated a highly patchy distribution of meiofauna taxa in the Lima estuary, revealed by the high OTU turnover even between sampling points only a few meters apart. Hence, eDNA-based meiofauna surveys require consideration of the necessary sampling effort on relatively small spatial scales, as well as an appreciation of the tidal level-induced variation of these communities.This study was funded by the project “The NextSea: Next generation monitoring of coastal ecosystems in a scenario of global change” (operação NORTE-01-0145-FEDER-000032), supported by Norte Portugal Regional Operational Programme (NORTE 2020), under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). Maria Fais and Sofia Duarte were supported, respectively by a PhD (SFRH/BD/113547/2015) and a post-doc fellowship (SFRH/BPD/109842/2015) from FCT. The authors would like to thank Prof. Jesús Troncoso (University of Vigo) and Prof. Pedro Gomes (University of Minho) for their availability and hospitality for the lab processing, as well as Barbara Leite (University of Minho) and Manuel Mendoza (University of Vigo) for their help during the sampling and data analysis phases, respectively