ArticleG3 (Bethesda, Md.)2017
Using Next-Generation Sequencing for DNA Barcoding: Capturing Allelic Variation in ITS2.
Article in G3 (Bethesda, Md.), 2017. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 19 papers.
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Who cites it
19 citing papers in PubMed, 59 citations in OpenAlex.
- Zoonotic malaria in Southeast Asia's changing landscapes: vector complexity, challenges, and future strategies.Parasites & vectors · 2026Review
- An All-in-One Metabarcoding Approach to Mosquito and Arbovirus Xenosurveillance.Molecular ecology resources · 2025Article
- Environmental Yeast Abundance and Diversity Assessment in Recreation Areas of Bangkok, Thailand.Environmental microbiology reports · 2025Article
- Genetic characterization and phylogenetic analysis of common house crows (Corvus splendens).Scientific reports · 2025Article
- Barcoding of Italian mosquitoes (BITMO): generation and validation of DNA barcoding reference libraries for native and alien species of Culicidae.Parasites & vectors · 2024Article
- Comparison of a multiplex PCR with DNA barcoding for identification of container breeding mosquito species.Parasites & vectors · 2024Article
- Determination of ITS1 haplotypes of Fritillariae Cirrhosae Bulbus by amplicon sequencing.Chinese medicine · 2024Article
- Long-term co-circulation of multiple arboviruses in southeast Australia revealed by xeno-monitoring and viral whole-genome sequencing.Virus evolution · 2024Article
- Article
- Application of next-generation sequencing to identify different pathogens.Frontiers in microbiology · 2023Review
- Challenges in Medicinal and Aromatic Plants DNA Barcoding-Lessons from the Lamiaceae.Plants (Basel, Switzerland) · 2022Review
- Using High-Throughput Amplicon Sequencing to Evaluate Intragenomic Variation and Accuracy in Species Identification ofJournal of fungi (Basel, Switzerland) · 2021Article
- Mosquitoes of the Maculipennis complex in Northern Italy.Scientific reports · 2021Article
- Article
- Advances in DNA Barcoding of Toxic Marine Organisms.International journal of molecular sciences · 2018Review
- Population genomics of Culiseta melanura, the principal vector of Eastern equine encephalitis virus in the United States.PLoS neglected tropical diseases · 2018Article
- Effective mosquito and arbovirus surveillance using metabarcoding.Molecular ecology resources · 2018Article
- An improved environmental DNA assay for bull trout (Salvelinus confluentus) based on the ribosomal internal transcribed spacer I.PloS one · 2018Article
- Assessing intragenomic variation of the internal transcribed spacer two: Adapting the Illumina metagenomics protocol.PloS one · 2017Article
Corrections and comments
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Authors and funding
4 authors at 2 institutions in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
Internal Transcribed Spacer 2 (ITS2) is a popular DNA barcoding marker; however, in some animal species it is hypervariable and therefore difficult to sequence with traditional methods. With next-generation sequencing (NGS) it is possible to sequence all gene variants despite the presence of single nucleotide polymorphisms (SNPs), insertions/deletions (indels), homopolymeric regions, and microsatellites. Our aim was to compare the performance of Sanger sequencing and NGS amplicon sequencing in characterizing ITS2 in 26 mosquito species represented by 88 samples. The suitability of ITS2 as a DNA barcoding marker for mosquitoes, and its allelic diversity in individuals and species, was also assessed. Compared to Sanger sequencing, NGS was able to characterize the ITS2 region to a greater extent, with resolution within and between individuals and species that was previously not possible. A total of 382 unique sequences (alleles) were generated from the 88 mosquito specimens, demonstrating the diversity present that has been overlooked by traditional sequencing methods. Multiple indels and microsatellites were present in the ITS2 alleles, which were often specific to species or genera, causing variation in sequence length. As a barcoding marker, ITS2 was able to separate all of the species, apart from members of the Culex pipiens complex, providing the same resolution as the commonly used Cytochrome Oxidase I (COI). The ability to cost-effectively sequence hypervariable markers makes NGS an invaluable tool with many applications in the DNA barcoding field, and provides insights into the limitations of previous studies and techniques.
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