real-time pcr (8)

6959334072?profile=RESIZE_400x The popularity of high value Chia and Quinoa "superfoods", makes them vulnerable to substitution and mislabelling. A real-time PCR (polymerase chain reaction)  (qPCR) method has been developed for the authentication of these two seeds.The developed method using chia- and quinoa specific primer-probe sets based on TaqMan technology was validated, and specificity, cross-reactivity, limit of detection, efficiency and robustness were determined. The method was successfully applied to twelve (chia) and seven (quinoa) commercial composite samples, indicating its suitability to authenticate these two foods.

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4167177917?profile=RESIZE_710xThe common cuttlefish (Sepia officinalis) is highly appreciated by consumers, and Spanish reseachers have developed a rapid assay to determine its authenticity. The method  is based on TaqMan real-time PCR (Polymerase Chain Reaction) method to amplify a 122 base pairs (bp) fragment of the mitochondrial COI (Cytochrome Oxidase I) region. Reference and commercial samples of S. officinalis showed a threshold cycle (Ct) mean of 14.40, while the rest of the species examined did not amplify. No cross-reactivity was detected with any other species. The assay differentiates S. officinalis from other species of the genus Sepia and other cephalopod species, and works for fresh, frozen, grilled, cooked and canned samples of Sepia spp. 

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3887798641?profile=RESIZE_710xThere is a need for accurate methods to quantify meat species in meat products to check QUID (quantitive ingredient delarations). Chinese researchers have evaluated a real-time PCR method based on species-specific primers and probes from the mitochondrial cytb (cytochrome b) gene fragment for the identification and quantification of beef ingredient in commercial meat products. The method was developed and calibrated using three mixed matrices (pork, donkey and sheep with known proportions of beef, respectively). Results showed that the primers and probes were highly specific for beef in meat products, and the absolute detection limit of the real-time PCR method was 0.025 ng DNA, and the relative detection limit was 0.002% (w/w) of beef.  The assay was validated with 22 commercial beef products, of which 11 were salted, 10 were jerkies (dried) and one meatball, which were collected from local supermarkets. The results indicated the assay had a good stability in detecting and quantifying beef in the commercial samples.  

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3817380870?profile=RESIZE_710xIn order to verify labelling compliance and evaluate the existence of fraudulent practices, 250 sausage samples were purchased from local markets in Sichuan Province and analysed for the presence of DNA from chicken, pork, beef, duck and genetically modified soybean using real-time and end-point PCR methods. In total, 74.4% (186) of the samples were properly labelled, while the other 25.6% (64) were mislabelled and potentially adulterated samples.The most common mislabelling was the undeclared addition of, or contamination with, duck meat, which is cheaper than pork or chicken.  

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Roumanian researchers have optimised and validated a real-time, sensitive, and accurate PCR method for the detection and quantification of meat species in selected processed meat products: chicken sausages, beef bologna, and pork bologna. A common detection limit of 8 DNA copies was established for each sample, corresponding to 0.1% w/w for beef and pork and 0.2% w/w for chicken. For the limit of quantification, dilutions of 20 copies of DNA for the bovine and pig species and 50 copies of DNA for the chicken species were performed. Specificity and selectivity tests in six replicates each showed no extraneous meat species, in line with the label information. Repeatability was assessed in six replicates, both quantitatively and qualitatively, by the same analyst, on the same day, and with the same equipment. The reproducibility results obtained by two analysts, on different days, for each sample were very similar. 

1337352593?profile=RESIZE_710x Read the full paper

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The aim of this project was to evaluate the applicability of selected modern molecular biology methods to reliably detect and quantify meat species around the 1% (w/w) level for enforcement action with a focus on processed meat products. Three techniques were evaluated, which had quantitative potential for trace ingredient detection: real-time PCR (qPCR), droplet digital PCR (ddPCR), and a label-free mass-spectrometry (MS) approach.

Results of the research indicated that both qPCR and ddPCR demonstrated good qualitative and quantitative analytical performance at the 1% (w/w) adulteration level for enforcement action (with an associated 3-27% coefficient of variation). Across all adulterant levels investigated, ddPCR generally showed tighter precision estimates, particularly at the 0.1% (w/w) levels and with the highly processed canned meat sample. The label-free MS technique demonstrated clear qualitative capability, but did not demonstrate comparable accuracy for quantitative determinations.  

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A real-time PCR-based screening assay was developed for the detection of crustaceans in food. In order to cover most relevant species in one analytical step, PCR systems were newly developed for the detection of prawns (Penaeidae), lobster (Homarus sp.), Common shrimp (Crangon crangon), river prawns (Macrobrachium sp.) and Chinese mitten crab (Eriocheir sinensis). In addition, a published system targeting Northern prawn (Pandalus borealis) was selected. All PCR-systems are based on mitochondrial 16S rRNA gene sequences, and were optimised to be run with a standard programme at a universal annealing temperature of 60 °C. After validation, the assay was tested on a range of food products. The assay enables detection of multiple species of market relevance.

Read the abstract at: RT-PCR Detection of Crustaceans

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species-specific PCR and real-time PCR with EvaGreen dye targeting the ITS region of Carthamus tinctorius L. (safflower) were successfully developed. A normalised real-time PCR approach was also proposed in the range of 0.1–20% (w/w) of safflower in saffron, which was successfully validated and applied to commercial saffron samples (stigmas, powders and seasonings).

Read the abstract at: Safflower adulteration of saffron

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