COI barcode-HRM as a novel approach for the discrimination of hake species

Fernandes, Telmo J.R.; Costa, Joana; Oliveira, M. Beatriz P.P.; Mafra, Isabel

doi:10.1016/j.fishres.2017.09.014

Cited by 30 publications

(26 citation statements)

References 37 publications

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“…All tissue samples from outside of the United States (e.g., Trinidad and Tobago, St. Lucia, Brazil) were initially preserved in ethanol, but were shipped wrapped in ethanol-soaked paper towels at 10 • C-25 • C, which may have compromised tissue preservation during shipping to our lab in Baltimore (United States), reducing the quality of extracted DNA. A number of other studies using HRM analysis for individual SNP genotyping have noted that DNA quality can greatly impact the quality and reliability of HRM genotyping (e.g., Garritano et al, 2009, but see Fitzcharles, 2012;Fernandes et al, 2018). It is possible that variation in the initial quality of DNA extractions may have increased the intra-species variation in melt profiles, reducing discriminatory power of the LDA models.…”

Section: Hrm-based Identification Of Callinectes Speciesmentioning

confidence: 99%

“…Combining the power of HRM with clustering statistics or supervised machine learning approaches allows further discrimination of highly similar sequences in a probabilistic framework (Reja et al, 2010;Winder et al, 2011;Bowman et al, 2017). HRM is beginning to be used for identification of marine species in the seafood industry (Fitzcharles, 2012;Jin et al, 2015;Fernandes et al, 2018;Verrez-Bagnis et al, 2018). Blue crab researchers, as well as the fisheries industry, could benefit from a rapid and cost-effective method for reliable identification of species within the Callinectes genus.…”

Section: Introductionmentioning

confidence: 99%

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Rapid Genetic Identification of the Blue Crab Callinectes sapidus and Other Callinectes spp. Using Restriction Enzyme Digestion and High Resolution Melt (HRM) Assays

et al. 2020

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The blue crab Callinectes sapidus is one of the most widely studied marine crustaceans due to its high economic value and ecological significance. Despite extensive research on the blue crab in North America, many questions remain about the distribution and abundance of the species in the subtropics and tropics. In many places, C. sapidus is sympatric with morphologically similar Callinectes spp., which has implications for seafood mislabeling. To enable rapid identification of the species, we designed and tested two PCR-based assays targeting the 12S rRNA mitochondrial gene. The first assay discriminates C. sapidus from other Callinectes spp. via post-PCR restriction digestion (PCR-RFLP) and the second assay discriminates among multiple Callinectes spp. through High Resolution Melting (HRM) analysis and supervised machine learning analyses. A total of 58 DNA samples from five Callinectes spp. (validated via 12S gene sequencing) were used for assay testing. The PCR RFLP assay was 100% accurate identifying C. sapidus from other Callinectes spp. HRM analysis of amplicons showed good discrimination among species, with distinct clusters formed between species with higher sequence homology. Linear discriminant analysis (LDA) classification of HRM curves was quite successful given the small dataset available, producing ∼90-91% mean accuracy in classification over all species with 100-fold cross validation. Much of the error came from misclassifications between C. similis and C. danae, which are ∼99% similar in sequence for the amplicon; collapsing them into a single class increased overall classification success to 94%. Error also arose from C. bocourti classifications, which had a reference set containing only three samples. Classification accuracy of C. sapidus alone via HRM was 97.5%. Overall, these assays show great promise as rapid and inexpensive methods to identify Callinectes spp. and have application for both ecological research and seafood identification or labeling.

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Section: Hrm-based Identification Of Callinectes Speciesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Rapid Genetic Identification of the Blue Crab Callinectes sapidus and Other Callinectes spp. Using Restriction Enzyme Digestion and High Resolution Melt (HRM) Assays

et al. 2020

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“…By comparing the melting curves of unknown samples, i.e. the change in intensity of the fluorescence signal with increasing temperature, with profiles of reliably identified samples, they can be assigned to known species [ 10 , 11 ].…”

Section: Introductionmentioning

confidence: 99%

Discriminating larvae of two syntopic Cychramus species (Coleoptera, Nitidulidae) by means of bar-HRM analysis

Zangl

Oberreiter

Huss³

et al. 2020

Mol Biol Rep

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Molecular genetic methods are increasingly used to supplement or substitute classical morphology-based species identification. Here, we employ a COI mini-barcode coupled high-resolution melting analysis to quickly, cost-efficiently and reliably determine larvae of two closely related Cychramus (Coleoptera, Nitidulidae) species. Euclidean distance comparison (p < 0.01) and a Welch t-test of the melting point temperatures (p < 0.01) provide highly significant statistical evidence for species specific differences in melting and fluorescence curves, thus allowing the assignment of larvae to either of the two species. This protocol serves as a fast, low-cost and low-tech method to discriminate between pairs or groups of closely related species and can be adapted and applied to various ecological research questions.

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“…However, in many circumstances, recovering sequences is not possible because only degraded DNA is available (Boyer et al 2012), which is the major problem of specimens older than a decade (Hajibabaei et al 2006). The High-Resolution Melting (HRM) analysis is an alternative method to sequencing to overcome this problem (Fernandes et al 2017;Osathanunkul et al 2018). This is a RT-PCR-based and non-contaminated post-PCR technique that allows analyzing genetic variation in small PCR amplicons (usually 80-120 bp) by detecting small differences in the melting temperatures of the sequences through the melting curves (Wittwer 2009;Wittwer et al 2003).…”

Section: Introductionmentioning

confidence: 99%

Peer Review #1 of "Using high-resolution melting to identify Calliphoridae (blowflies) species from Brazil (v0.1)"

Malewski¹

2020

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COI barcode-HRM as a novel approach for the discrimination of hake species

Cited by 30 publications

References 37 publications

Rapid Genetic Identification of the Blue Crab Callinectes sapidus and Other Callinectes spp. Using Restriction Enzyme Digestion and High Resolution Melt (HRM) Assays

Rapid Genetic Identification of the Blue Crab Callinectes sapidus and Other Callinectes spp. Using Restriction Enzyme Digestion and High Resolution Melt (HRM) Assays

Discriminating larvae of two syntopic Cychramus species (Coleoptera, Nitidulidae) by means of bar-HRM analysis

Peer Review #1 of "Using high-resolution melting to identify Calliphoridae (blowflies) species from Brazil (v0.1)"

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