Droplet digital PCR (ddPCR) method for the detection and quantification of goat and sheep derivatives in commercial meat products

Wang, Qiang; Cai, Yicun; He, Yihua; Liu, Yang; Li, Jian; Pan, Liangwen

doi:10.1007/s00217-017-3000-5

Cited by 28 publications

(18 citation statements)

References 27 publications

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“…4 dPCR has been widely used in many areas such as food authentication, identification of genetically modified organisms, and clinical testing. 4,6,17 We established a method for detection and quantification of PEDV using droplet digital PCR (ddPCR).…”

mentioning

confidence: 99%

Development of a droplet digital PCR for detection and quantification of porcine epidemic diarrhea virus

Cao

Chen

et al. 2020

J VET Diagn Invest

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Porcine epidemic diarrhea, a disease caused by porcine epidemic diarrhea virus (PEDV), results in large economic losses to the global swine industry. To manage this disease effectively, it is essential to detect PEDV early and accurately. We developed a sensitive and accurate droplet digital PCR (ddPCR) assay to detect PEDV. The optimal primer-to-probe concentration and melting temperature were identified as 300:200 nM and 59.2°C, respectively. The specificity of the ddPCR assay was confirmed by negative test results for common swine pathogens. The detection limit for the ddPCR was 0.26 copies/μL, which is a 5.7-fold increase in sensitivity compared to that of real-time PCR (rtPCR). Both ddPCR and rtPCR assays exhibited good linearity, although ddPCR provided higher sensitivity for clinical detection compared to that of rtPCR. Our ddPCR methodology provides a promising tool for evaluating the PEDV viral load when used for clinical testing, particularly for detecting samples with low-copy viral loads.

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mentioning

confidence: 99%

Development of a droplet digital PCR for detection and quantification of porcine epidemic diarrhea virus

Cao

Chen

et al. 2020

J VET Diagn Invest

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“…Reference DNA from other species, purchased from Zyagen Laboratories (San Diego, CA, USA), was used to verify the specificity of the assays. Since muscle tissue is the most common animal tissue used for cooking or as a component material of other products [ 27 ], porcine and chicken muscle tissue was used to develop the analytical methods in this study. Tissue samples were minced, dried in a baking oven (UFE500AO, Memeert, Germany) at 80°C for 72 h, then milled into superfine powder in liquid nitrogen using a Freezer Mixer (6850 freezer/mill, SPEX Sample Prep, USA).…”

Section: Methodsmentioning

confidence: 99%

“…The specificity of the methods, including assay exclusivity and inclusivity, were determined according to established practices [ 27 , 31 ]. To assess exclusivity, DNA samples from porcine ( S .…”

Section: Methodsmentioning

confidence: 99%

Collaborative ring trial of two real-time PCR assays for the detection of porcine- and chicken-derived material in meat products

Wang

Cai

et al. 2018

PLoS ONE

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In this study, we describe an inter-laboratory collaborative ring trial validation of species-specific TaqMan real-time PCR assays for the detection of porcine- and chicken-derived materials in meat products. We comprehensively evaluated the performance of these assays in different environments and situations. This validation included the participation of thirteen laboratories across Europe and Asia. The results from the thirteen participating laboratories were analyzed to determine the specificity, accuracy, false positive rate, limit of detection (LOD), and probability of detection (POD) of the developed methods. These results indicated that the methods developed to detect porcine- and chicken-derived materials in meat products are robust and repeatable. The false positive and false negative rates were both 0%, and the LOD was determined to be five copies/reaction. The laboratory standard deviation (σL) was 0.30 for both detection methods, indicating that the developed methods are suitable for detection and identification of the porcine- and chicken-derived materials in meat products.

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“…Methods such as qPCR and dPCR have been gaining popularity to quantify the amount of target and unknown adulterants 18 in food and feed samples 19 , meat products 20 , 21 , fish products 22 , microbial ecology 23 , GMOs 24 . This is the first study to quantify botanicals using qPCR and dPCR methods.…”

Section: Introductionmentioning

confidence: 99%

Quantification of Actaea racemosa L. (black cohosh) from some of its potential adulterants using qPCR and dPCR methods

Shanmughanandhan

Dhivya

Ragupathy

et al. 2021

Sci Rep

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The demand for popular natural health products (NHPs) such as Black Cohosh is increasing considerably, which in turn challenges quality assurance (QA) throughout the supply chain. To detect and quantify the target species present in a given NHP, DNA-based molecular techniques such as Real-time quantitative PCR (qPCR) and digital PCR (dPCR) are standard tools in the food and pathogen testing industries. There is a gap in the literature concerning validated quantitative PCR methods for botanicals that can be utilized for QA and good manufacturing practices. The objective of this study is to develop an efficient quantification method using qPCR and dPCR techniques for the detection and quantification of Actaea racemosa (Black cohosh) NHPs from its potential adulterants. These developed methods are validated for applicability on commercial NHPs. Species-specific hydrolysis probe assays were designed to analyze the black cohosh NHPs using qPCR and dPCR techniques. The results confirmed that the developed qPCR and dPCR methods are highly precise for identifying and quantifying black cohosh NHPs, indicating their potential applicability in future routine industrial and laboratory testing. This enables a single qPCR test to determine not only the presence of a specific botanical, but also the amount when mixed with an adulterant.

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Droplet digital PCR (ddPCR) method for the detection and quantification of goat and sheep derivatives in commercial meat products

Cited by 28 publications

References 27 publications

Development of a droplet digital PCR for detection and quantification of porcine epidemic diarrhea virus

Development of a droplet digital PCR for detection and quantification of porcine epidemic diarrhea virus

Collaborative ring trial of two real-time PCR assays for the detection of porcine- and chicken-derived material in meat products

Quantification of Actaea racemosa L. (black cohosh) from some of its potential adulterants using qPCR and dPCR methods

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