Development of a COX 1 based PCR-RFLP method for fish species identification

Mueller, Steffen; Handy, Sara M.; Deeds, Jonathan R.; George, Gideon O.; Broadhead, Wendy J.; Pugh, Sian E.; Garrett, S. D.

doi:10.1016/j.foodcont.2015.02.026

Cited by 30 publications

(16 citation statements)

References 15 publications

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“…; Mueller et al . ). Similarly, species‐specific real‐time PCR systems have been designed for tephritid fruit flies based on COI barcode libraries (Dhami et al .…”

Section: Discussionmentioning

confidence: 97%

A transcriptome‐based analytical workflow for identifying loci for species diagnosis: a case study with Bactrocera fruit flies (Diptera: Tephritidae)

et al. 2017

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Development of novel molecular methods for accurate and economical identification of species has become critical both for pure biological research and for a wide range of applied areas. The most widely used current molecular diagnostic tool, the mitochondrial cytochrome c oxidase subunit 1 gene (COI), the so-called DNA barcode, has been highly criticised and is known to be ineffective at distinguishing species in many groups. Alternative markers are needed to circumvent these issues and provide diagnosticians with a greater range of tools for making accurate identifications. To address this, we describe here a novel analytical workflow for diagnostic marker development that utilises near-genomic-scale data to search for potential informative loci. The workflow takes advantage of orthologous gene databases, in combination with tests of phylogenetic resolution, and benchmarking of nucleotide variation against COI, to determine putative loci that might outperform COI. We use transcriptomes of 14 tephritid fruit flies and especially the taxonomically complex genus Bactrocera, as a case study. Of 1646 orthologues searched, our workflow retained a total of five loci following our conservative filtering strategy. One locus, POP4, had strong potential as a novel diagnostic marker for Bactrocera fruit flies. POP4 discriminates most species in the training set of taxa, but like COI fails to separate the sibling species Bactrocera tryoni and Bactrocera neohumeralis. Further validation of this potential new marker against a broader taxonomic sample is ongoing. We advocate that this simple and efficient workflow is, with minor modification, customisable for diagnostic development in almost any taxonomic group.

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“…; Mueller et al . ). Similarly, species‐specific real‐time PCR systems have been designed for tephritid fruit flies based on COI barcode libraries (Dhami et al .…”

Section: Discussionmentioning

confidence: 97%

A transcriptome‐based analytical workflow for identifying loci for species diagnosis: a case study with Bactrocera fruit flies (Diptera: Tephritidae)

et al. 2017

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“…The mitochondrial COI gene is used most frequently for DNA-based species identification (Soroka, 2008). Mueller et al, (2015) developed a fast complementary method for the generation of PCR-RFLP patterns using a novel 439 bp amplicon within the established COX1 barcoding gene fragment that contains restriction sites for the fast restriction enzymes MluCI and HpaII.…”

Section: Discussionmentioning

confidence: 99%

Untitled

Kalaycı¹,

Akhan²

2016

Turk. J. Fish. Aquat. Sci.

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AbstractmtDNA-RFLP, a technique commonly used for molecular identification of species, was used to distinguish between morphologically similar crayfish species Austropotamobius torrentium and Astacus leptodactylus in this study. The mitochondrial COI gene was amplified and digested using 15 restriction endonucleases to establish a restriction map. The COI gene sequence contained recognition sites for three of the restriction endonucleases (SspI, KpnI, and RsaI) and produced two fragments and three genotypes for Austropotamobius torrentium. However, the Astacus leptodactylus COI gene did not have restriction sites for these three enzymes. Thus, a mtDNA-RFLP using these three endonucleases can be used to successfully distinguish between these two species of crayfish.Keywords: crayfish, species identification, mtDNA, COI. Astacus leptodactylus ve Austropotamobius torrentium Türlerinin mtDNA-RFLP Metodu ile Moleküler Teşhisi ÖzetBu çalışmada, canlı türlerinin moleküler olarak teşhisinde yaygın olarak kullanılan mtDNA-RFLP tekniği, morfolojik olarak birbirine benzeyen Austropotamobius torrentium and Astacus leptodactylus kervit türlerinin ayrımında kullanılmıştır. Mitokondrial COI gen bölgesi polimeraz zincir reaksiyonu ile çoğaltılarak 15 farklı endonükleaz enzimi ile kesilmiştir. Bu kesim enzimlerinden üç tanesi (SspI, KpnI, and RsaI) çoğaltılan COI gen bölgesini keserek Austropotamobius torrentium türü için iki parçacık ve üç genotip üretirken, Astacus leptodactylus türü için kesim bölgesi bulamamıştır. Çalışma sonuçları bu üç kesim enziminin mtDNA-RFLP tekniği kullanılarak bu iki kerevit türün ayrımında başarılı bir şekilde kullanılabileceğini göstermiştir.

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“…Conserved genomic sequences, referred to as DNA barcodes, are remarkably useful targets in genomic food analyses . A barcode sequence can be amplified simultaneously from all species constituents in a sample.…”

Section: Introductionmentioning

confidence: 99%

The trnL (UAA)‐trnF (GAA) intergenic spacer is a robust marker of green pea (Pisum sativum L.) adulteration in economically valuable pistachio nuts (Pistacia vera L.)

Sen

Uncu

et al. 2020

J Sci Food Agric

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BACKGROUND Pistachio (Pistacia vera L.) is an expensive culinary nut species; it is therefore susceptible to adulteration for economic profit. Green pea (Pisum sativum L.) kernels constitute the most common material used for adulterating chopped / ground pistachio nuts and pistachio paste. Food genomics enables the species composition of a food sample to be ascertained through DNA analysis. Accordingly, a barcode DNA genotyping approach was used to standardize a test method to identify green pea adulteration in pistachio nuts. RESULTS The trnL (UAA)‐trnF (GAA) intergenic spacer in the plastid genome was the target analyte in the present study. The barcode locus displayed a significant, discriminatory size difference between pistachio and pea, with amplicon sizes of 449 and 179 bp, respectively. Polymerase chain reaction‐capillary electrophoresis (PCR‐CE) analysis of the intergenic spacer resulted in the successful identification of species composition in the in‐house admixtures, which contained 5% to 30% of green pea. CONCLUSION The present work describes a fast and straightforward DNA test that identifies green pea adulteration in pistachio nuts without requiring a statistical data interpretation process. The plastid trnL (UAA)‐trnF (GAA) intergenic spacer length widely varies among plant taxa, so the PCR‐CE protocol that operates on the intergenic spacer holds the potential to reveal adulteration with a plethora of adulterants. The PCR‐CE assay described in the present work can be adopted readily by food‐quality laboratories in the public sector or the food industry as an easy and reliable method to analyze pistachio authenticity. © 2020 Society of Chemical Industry

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Development of a COX 1 based PCR-RFLP method for fish species identification

Cited by 30 publications

References 15 publications

A transcriptome‐based analytical workflow for identifying loci for species diagnosis: a case study with Bactrocera fruit flies (Diptera: Tephritidae)

A transcriptome‐based analytical workflow for identifying loci for species diagnosis: a case study with Bactrocera fruit flies (Diptera: Tephritidae)

Untitled

The trnL (UAA)‐trnF (GAA) intergenic spacer is a robust marker of green pea (Pisum sativum L.) adulteration in economically valuable pistachio nuts (Pistacia vera L.)

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