Dataset Information

Method for quick DNA barcode reference library construction.

ABSTRACT: DNA barcoding has become one of the most important techniques in plant species identification. Successful application of this technology is dependent on the availability of reference database of high species coverage. Unfortunately, there are experimental and data processing challenges to construct such a library within a short time. Here, we present our solutions to these challenges. We sequenced six conventional DNA barcode fragments (ITS1, ITS2, matK1, matK2, rbcL1, and rbcL2) of 380 flowering plants on next-generation sequencing (NGS) platforms (Illumina Hiseq 2500 and Ion Torrent S5) and the Sanger sequencing platform. After comparing the sequencing depths, read lengths, base qualities, and base accuracies, we conclude that Illumina Hiseq2500 PE250 run is suitable for conventional DNA barcoding. We developed a new "Cotu" method to create consensus sequences from NGS reads for longer output sequences and more reliable bases than the other three methods. Step-by-step instructions to our method are provided. By using high-throughput machines (PCR and NGS), labeling PCR, and the Cotu method, it is possible to significantly reduce the cost and labor investments for DNA barcoding. A regional or even global DNA barcoding reference library with high species coverage is likely to be constructed in a few years.

SUBMITTER: Liu Y

PROVIDER: S-EPMC8427591 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Method for quick DNA barcode reference library construction.

Liu Yanlei Y Xu Chao C Sun Yuzhe Y Chen Xun X Dong Wenpan W Yang Xueying X Zhou Shiliang S

Ecology and evolution 20210804 17

DNA barcoding has become one of the most important techniques in plant species identification. Successful application of this technology is dependent on the availability of reference database of high species coverage. Unfortunately, there are experimental and data processing challenges to construct such a library within a short time. Here, we present our solutions to these challenges. We sequenced six conventional DNA barcode fragments (ITS1, ITS2, matK1, matK2, rbcL1, and < ...[more]

PMID: 34522329

Similar Datasets

Project description:In the last few years, DNA barcoding became an established method for species identification in biodiversity inventories and monitoring studies. Such studies depend on the access to a comprehensive reference data base, covering all relevant taxa. Here we present a comprehensive DNA barcode inventory of all amphibian and reptile species native to Austria, except for the putatively extinct Vipera ursinii rakosiensis and Lissotriton helveticus, which has been only recently reported for the very western edge of Austria. A total of 194 DNA barcodes were generated in the framework of the Austrian Barcode of Life (ABOL) initiative. Species identification via DNA barcodes was successful for most species, except for the hybridogenetic species complex of water frogs (Pelophylax spp.) and the crested newts (Triturus spp.), in areas of sympatry. However, DNA barcoding also proved powerful in detecting deep conspecific lineages, e.g. within Natrix natrix or the wall lizard (Podarcis muralis), resulting in more than one Barcode Index Number (BIN) per species. Moreover, DNA barcodes revealed the presence of Natrix helvetica, which has been elevated to species level only recently, and genetic signatures of the Italian water frog Pelophylax bergeri in Western Austria for the first time. Comparison to previously published DNA barcoding data of European amphibians and reptiles corroborated the results of the Austrian data but also revealed certain peculiarities, underlining the particular strengths and in the case of the genus Pelophylax also the limitations of DNA barcoding. Consequently, DNA barcoding is not only powerful for species identification of all life stages of most Austrian amphibian and reptile species, but also for the detection of new species, the monitoring of gene flow or the presence of alien populations and/or species. Thus, DNA barcoding and the data generated in this study may serve both scientific and national or even transnational conservation purposes.

Project description:BackgroundAmomum villosum Lour. is the plant that produces the famous traditional Chinese medicine Amomi Fructus. Frequent habitat destruction seriously threatens A. villosum germplasm resources. Genetic diversity is very important to the optimization of germplasm resources and population protection, but the range of inherited traits within A. villosum is unclear. In this study, we analyzed the genetic diversity and genetic structures of A. villosum populations in Guangdong and constructed a local reference DNA barcode library as a resource for conservation efforts.MethodsDNA barcoding and Inter-Simple Sequence Repeat (ISSR) markers were used to investigate the population genetics of A. villosum. Five universal DNA barcodes were amplified and used in the construction of a DNA barcode reference library. Parameters including percentage of polymorphic sites (PPB), number of alleles (Na), effective number of alleles (Ne), Nei's gene diversity index (H), and Shannon's polymorphism information index (I) were calculated for the assessment of genetic diversity. Genetic structure was revealed by measuring Nei's gene differentiation coefficient (Gst), total population genetic diversity (Ht), intra-group genetic diversity (Hs), and gene flow (Nm). Analysis of molecular variance (AMOVA), Mantel tests, unweighted pair-group method with arithmetic mean (UPGMA) dendrogram, and principal co-ordinates (PCoA) analysis were used to elucidate the genetic differentiation and relationship among populations.ResultsA total of 531 sequences were obtained from the five DNA barcodes with no variable sites from any of the barcode sequences. A total of 66 ISSR bands were generated from A. villosum populations using the selected six ISSR primers; 56 bands, 84.85% for all the seven A. villosum populations were polymorphic. The A. villosum populations showed high genetic diversity (H = 0.3281, I = 0.4895), whereas the gene flow was weak (Nm = 0.6143). Gst (0.4487) and AMOVA analysis indicated that there is obvious genetic differentiation amongA. villosum populations and more genetic variations existed within each population. The genetic relationship of each population was relatively close as the genetic distances were between 0.0844 and 0.3347.

Dataset Information

Method for quick DNA barcode reference library construction.

Publications

Method for quick DNA barcode reference library construction.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets