A new method for the rapid detection of Atlantic cod (Gadus morhua), Pacific cod (Gadus macrocephalus), Alaska pollock (Gadus chalcogrammus) and ling (Molva molva) using a lateral flow dipstick assay.

Species-specific lateral flow dipstick (LFD) assays for the identification of Atlantic cod (Gadus morhua), Pacific cod (Gadus macrocephalus), Alaska pollock (Gadus chalcogrammus) and ling (Molva molva) in food products were developed. The method comprises a PCR system with four sets of specific primers, for each target species. This step was also devised to dual-labeling of PCR products with biotin and 6-FAM, which are then easily read on a lateral flow dipstick, upon which these products are immobilized by a fixed biotin-ligand and visualized with anti-FAM antibody-coated gold nanoparticles. Sensitivity and selectivity were determined for each of the developed assays. Validation of the assays was performed with DNA extracted from commercial fish products, the identification of all samples by PCR-LFD was coherent with the results found with DNA sequencing. Target species were successfully detected in analyzed commercial samples, demonstrating the applicability of this method to the rapid analysis of food products.

A new real-time PCR method for rapid and specific detection of ling (Molva molva).

Seafood fraud - often involving substitution of one species by another - has attracted much attention as it is prevalent worldwide. Whilst DNA analysis has helped to combat this type of fraud some of the methods currently in use are time-consuming and require sophisticated equipment or highly-trained personnel. This work describes the development of a new, real-time PCR TaqMan assay for the detection of ling (Molva molva) in seafood products. For this purpose, specific primers and a minor groove binding (MGB) TaqMan probe were designed to amplify the 81bp region on the cyt b gene. Efficiency, specificity and cross-reactivity assays showed statistically significant differences between the average Ct value obtained for Molva molva DNA (19.45±0.65) and the average Ct for non-target species DNA (38.3±2.8), even with closely related species such as Molva dypterygia (34.9±0.09). The proposed methodology has been validated with 31 commercial samples.

Identification of Atlantic cod (Gadus morhua), ling (Molva molva), and Alaska pollock (Gadus chalcogrammus) by PCR-ELISA using duplex PCR.

Species-specific PCR-ELISA assays for the identification of Atlantic cod (Gadus morhua), Alaska pollock (Gadus chalcogrammus), and ling (Molva molva) in food products have been developed. The method, comprising a set of primers common to the first two species, a set of primers for M. molva, and a probe for each species, was designed using ND4 and cytochrome b genes as molecular markers. The sensitivity and selectivity were then determined for each assay. These assays were afterward used to analyze DNA extracted from commercial fish products. The presence of the target species was successfully detected in all analyzed samples, demonstrating the applicability of this method to the analysis of food products.