Development of a real-time PCR system for the detection of the potential allergen fish in food

Tetzlaff, Carina; Mäde, Dietrich

doi:10.1007/s00217-016-2799-5

Cited by 22 publications

(11 citation statements)

References 11 publications

(16 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Then, 10 µL RNase A [c = 30 mg/mL] (Macherey-Nagel, Düren, Germany) was added and incubated again for 45 min at 65 °C with shaking. Further workup was performed as described in [32]. In brief, 10 µL Proteinase K solution (c = 20 mg/L) was added, and the samples were incubated at 60 °C under agitation overnight and centrifuged for 10 min at 12,000×g afterwards.…”

Section: Sample Preparation and Dna Extractionmentioning

confidence: 99%

“…Sequencing of the purified PCR products was performed with the BigDye™ Terminator V 1.1 Cycle Sequencing Kit (Applied Biosystems, Darmstadt, Germany) using the same primers as those in conventional PCR. The reaction conditions of the sequence reaction were chosen analogously to [32]. The annealing temperature of the primers was 5 °C below the melting temperature; see Electronic Supplementary Material (Supp.…”

Section: Dna-sequencingmentioning

confidence: 99%

See 1 more Smart Citation

Development and validation of a method for quantification of common wheat, durum wheat, rye and barley by droplet digital PCR

Schulze

Geuthner

Mäde

2021

Eur Food Res Technol

Self Cite

View full text Add to dashboard Cite

Food fraud is becoming a prominent topic in the food industry. Thus, valid methods for detecting potential adulterations are necessary to identify instances of food fraud in cereal products, a significant component of human diet. In this work, primer–probe systems for real-time PCR and droplet digital PCR (ddPCR) for the detection of these cereal species: bread wheat (together with spelt), durum wheat, rye and barley for real-time PCR and ddPCR were established, optimized and validated. In addition, it was projected to validate a molecular system for differentiation of bread wheat and spelt; however, attempts for molecular differentiation between common wheat and spelt based on the gene GAG56D failed because of the genetic variability of the molecular target. Primer–probe systems were further developed and optimized on the basis of alignments of DNA sequences, as well as already developed PCR systems. The specificity of each system was demonstrated on 10 (spelt), 11 (durum wheat and rye) and 12 (bread wheat) reference samples. Specificity of the barley system was already proved in previous work. The calculated limits of detection (LOD95%) were between 2.43 and 4.07 single genome copies in real-time PCR. Based on the “three droplet rule”, the LOD95% in ddPCR was calculated to be 9.07–13.26 single genome copies. The systems were tested in mixtures of flours (rye and common wheat) and of semolina (durum and common wheat). The methods proved to be robust with regard to the tested conditions in the ddPCR. The developed primer–probe systems for ddPCR proved to be effective in quantitatively detecting the investigated cereal species rye and common wheat in mixtures by taking into account the haploid genome weight and the degree of milling of a flour. This method can correctly detect proportions of 50%, 60% and 90% wholemeal rye flour in a mixture of wholemeal common wheat flour. Quantitative results depend on the DNA content, on ploidy of cereal species and are also influenced by comminution. Hence, the proportion of less processed rye is overestimated in higher processed bread wheat and adulteration of durum wheat by common wheat by 1–5% resulted in underestimation of common wheat.

show abstract

Section: Sample Preparation and Dna Extractionmentioning

confidence: 99%

Section: Dna-sequencingmentioning

confidence: 99%

Development and validation of a method for quantification of common wheat, durum wheat, rye and barley by droplet digital PCR

Schulze

Geuthner

Mäde

2021

Eur Food Res Technol

Self Cite

View full text Add to dashboard Cite

show abstract

“…Besides, another qPCR system was developed to trace crustaceans through targeting 16S rRNA mitochondrial gene in foods, allowing the Linacero et al, 2016) detection and quantification down to 0.1 pg and 0.0001% (w/w) of shrimp DNA and shrimp in model mixtures (Fernandes, Joana, Oliveira, & Isabel, 2018). Tetzlaff et al (2016) reported a qPCR method with an ability to indicate the presence of organisms belonging to Teleostei in food. A cost-effective qPCR scheme was established by Herrero, Vieites, and Espineira (2014) to confirm the presence of fish.…”

Section: Detection Of Food Allergens By Qpcrmentioning

confidence: 99%

“…Tetzlaff et al. (2016) reported a qPCR method with an ability to indicate the presence of organisms belonging to Teleostei in food. A cost‐effective qPCR scheme was established by Herrero, Vieites, and Espineira (2014) to confirm the presence of fish.…”

Section: Real‐time Fluorescence Quantification Pcrmentioning

confidence: 99%

Fluorescence‐based quantitative platform for ultrasensitive food allergen detection: From immunoassays to DNA sensors

Qian

Zhou

et al. 2020

Comp Rev Food Sci Food Safe

View full text Add to dashboard Cite

Food allergies are global health issue with an increasing prevalence that affect food safety; hence, food allergen detection, labeling, and management are considered to be important priorities in the food industry. In this critical review, we provide a comprehensive overview of several fluorescence-based platforms based on different biorecognition ligands, such as antibodies, DNA, aptamers, and cells, for food allergen quantification. Traditional analytical methods are generally unsuitable for food manufacturers to accomplish the real-time identification of food allergens in food products. Therefore, it is important to develop simple, rapid, inexpensive, accurate, and sensitive methods to improve user accessibility. A fluorescence-based quantitative platform provides an excellent detection platform for food allergens because of its high sensitivity. This review summarizes the traditional antibody-based fluorescent techniques for food allergen detection, such as the time-resolved fluoroimmunoassay , immunofluorescence imaging, fluorescence enzyme-linked immune sorbent assay, flow injection fluoroimmunoassay, and fluorescence immunosensors. However, these methods suffer from disadvantages such as the significant rate of false-positive and false-negative results due to antibody cross-reactivity with nontarget food components in the complex food matrix and epitope degradation during food processing. Hence, different types of fluorescence-based immunoassays are suitable for standardization and quantification of allergens in fresh foods. In addition, we summarize new fluorescence-based quantitative platforms, including fluorescence genosensors, fluorescence cell sensors, and fluorescence aptamer sensors. With the advantages of high sensitivity and simple operation, fluorescence biosensors will have great potential in the future and could provide portable methods for multiallergen real-time detection in complex food systems.

show abstract

“…While it does not detect allergenic proteins directly, PCR has proven to be more robust than protein‐based detection methods in cases of food matrix interference and food processing . Detection of various fish species has been carried out using PCR‐based assays for several targets, including the nuclear genes for parvalbumin, rhodopsin, Hoxc13, and 18S rRNA as well as the mitochondrial genes for 12S rRNA and 16S rRNA . Design of PCR‐based detection assays depends on the availability of relevant DNA sequence data from the species of interest to ensure accuracy and, ideally, incorporates sequences from related nontarget species to help ensure that cross‐reactivity does not occur.…”

Section: Introductionmentioning

confidence: 99%

A method to detect allergenic fish, specifically cod and pollock, using quantitative real‐time PCR and COI DNA barcoding sequences

Eischeid

2018

J Sci Food Agric

View full text Add to dashboard Cite

BACKGROUND: Fish are one of eight major allergens defined in the US Food Allergen Labeling and Consumer Protection Act, and cod and pollock are two of the major fish allergens. This paper describes development and validation of a method to detect cod and pollock in complex food matrices using real-time polymerase chain reaction (PCR). Mitochondrial cytochrome oxidase I (COI) sequences obtained through DNA barcoding were used to design a single set of primers and probe which detected three species in the genus Gadus: Atlantic cod, Pacific cod, and walleye pollock. RESULTS: Cod spiked into three different food matrices (cooking oil, clam chowder, and hushpuppy mix) yielded high linearity, dynamic range spanning six orders of magnitude, and lower limits of detection at 1-10 ppm (ppm; mg kg −1 ). Frying had an adverse effect on the lower limit of detection, but not on linearity.CONCLUSIONS: This work shows that COI DNA barcoding sequences can be used to effectively design real-time PCR assays for detection of food allergens in complex matrices. While full-length DNA barcodes distinguish individual species, the PCR assay designed here detected three different species. This is likely because real-time PCR assays are tolerant to basepair mismatches and do not utilize the full length of the DNA barcode. Published 2018. This article is a U.S. Government work and is in the public domain in the USA.

show abstract

Development of a real-time PCR system for the detection of the potential allergen fish in food

Cited by 22 publications

References 11 publications

Development and validation of a method for quantification of common wheat, durum wheat, rye and barley by droplet digital PCR

Development and validation of a method for quantification of common wheat, durum wheat, rye and barley by droplet digital PCR

Fluorescence‐based quantitative platform for ultrasensitive food allergen detection: From immunoassays to DNA sensors

A method to detect allergenic fish, specifically cod and pollock, using quantitative real‐time PCR and COI DNA barcoding sequences

Contact Info

Product

Resources

About