Real-time Polymerase Chain Reaction for the Food Microbiologist: Technologies, Applications, and Limitations

Hanna, Scott E.; Connor, Christopher; Wang, Hua H.

doi:10.1111/j.1365-2621.2005.tb07149.x

Cited by 82 publications

(56 citation statements)

References 35 publications

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Section: Real-time Pcr (Quantitative Pcr / Qpcr)mentioning

confidence: 99%

“…Real-time PCR assays have been increasingly used for specific, sensitive and quantitatively detection of species specific DNA fragments in meat products. Due to its specificity and low detection limits, it is preferred to determine species of processed products (Cai et al, 2012;Hanna et al, 2005). Agriculture -Food Science and Technology, 5(5): 507-517, 2017 509 Cai et al (2012) represented two species-specific realtime PCR assay as simple, reliable and sensitive method for detection and quantification of bovine and porcine DNA in gelatin mixtures and capsules.…”

Section: Real-time Pcr (Quantitative Pcr / Qpcr)mentioning

confidence: 99%

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Origin Determination and Differentiation of Gelatin Species of Bovine, Porcine, and Piscine through Analytical Methods

Eryılmaz

Işık

Demircan

et al. 2017

Turkish JAF Sci.Tech.

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Gelatin origin determination has been a crucial issue with respect to religion and health concerns. It is necessary to analyze the origin of gelatin with reliable methods to ensure not only consumer choices but also safety and legal requirements such as labeling. There are many analytical methods developed for detection and/or quantification of gelatin from different sources including bovine, porcine and piscine. These analytical methods can be divided into physicochemical, chromatographic, immunochemical, spectroscopic and molecular methods. Moreover, computational methods have been used in some cases consecutively to ensure sensitivity of the analytical methods. Every method has different advantages and limitations due to their own principles, applied food matrix and process conditions of material. The present review intends to give insight into novel analytical methods and perspectives that have been developed to differentiate porcine, bovine and piscine gelatins and to establish their authenticity. Almost every method can be succeeded in origin determination; however, it is a matter of sensitivity in that some researches fail to ensure sufficient differentiation. IntroductionGelatin is a water soluble protein obtained by hydrolysis of collagen to several high molecular weight polypeptides. While the main component of the gelatin is protein (85-92%), it also contains mineral salts and moisture (Baziwane and He, 2003;Schrieber and Gareis, 2007). Gelatin has a great importance in pharmaceutical, cosmetic and especially food industry with its well-known functional properties such as gel formation in jellies, foam formation and stabilization in ice cream, emulsion stabilization in meat products, syneresis stabilization in yogurt (Nhari et al., 2012). Gelatin was produced approximately 326.000 metric tons in 2008 worldwide, and it is estimated to be consumed 395.840 metric tons by the year of 2017 according to Global Industry Analysts, Inc. which is one of the largest and reputed market research companies (Anonymous, 2016).The raw materials used in gelatin production are mainly from bovine and porcine sources: 80% from pig skin, 15% from cattle hide split, the remaining 5 % from bones, fish and other sources (GME, 2016). Gelatin origin determination has been a matter of paramount importance for Muslims, Hindus, Jews, vegetarians and vegans, in that, in Islam and Judaism, porcine derivatives are prohibited to consume, whereas in Hinduism, cow can neither be slaughtered nor consumed. Vegetarians and vegans prefer to eat animal-free foods (Boran et al., 2010;Nhari et al., 2012). Moreover, bovine gelatin has been restricted for human consumption due to Bovine Spongiform Encephalopathy (BSE) outbreak in 1986 in England and other countries (Venien and Levieux, 2005). Therefore, it is necessary to analyze the origin of gelatin with a reliable method. As far as it is concerned; physiochemical, chromatographic, immunochemical, spectroscopic and molecular methods have been developed to determine and differentiate gelatin o...

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Section: Real-time Pcr (Quantitative Pcr / Qpcr)mentioning

confidence: 99%

Section: Real-time Pcr (Quantitative Pcr / Qpcr)mentioning

confidence: 99%

Origin Determination and Differentiation of Gelatin Species of Bovine, Porcine, and Piscine through Analytical Methods

Eryılmaz

Işık

Demircan

et al. 2017

Turkish JAF Sci.Tech.

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“…That is, when a food or clinical sample contains both dead and live target cells, the TaqMan real-time PCR detects both. This can be a serious problem in a food where processing may destroy microbial cells, but leave their DNA relatively intact 1) . On the other hand, enrichment of microorganisms in a sample with a nutrient broth has been recognized to be a useful procedure for subsequent real-time PCR 2)ῌ4) .…”

Section: Introductionmentioning

confidence: 99%

“…However, some problems remain even in the TaqMan real-time PCR. One of them is an inability to distinguish between live and dead cells 1) . That is, when a food or clinical sample contains both dead and live target cells, the TaqMan real-time PCR detects both.…”

Section: Introductionmentioning

confidence: 99%

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Estimation of Viable Salmonella Cell Numbers in Meat and Meat Product Using Real-Time PCR

Fujikawa¹,

Shimojima²

2008

J. Food Hyg. Soc. Jpn

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Recently we developed a novel method for estimating viable Salmonella cell numbers by means of a 5῎-nuclease real-time PCR [Fujikawa et al., J. Food Hyg. Japan, 47, 151ῌ156 (2006)]. The method was based on the increase kinetics of the target DNA region (inv A) of the microorganism growing in a culture medium during incubation. The index for the PCR was the threshold cycle. In this study, we validated the method for application in food. Namely, Salmonella cells spiked into ground chicken, pork, and beef and raw hamburger patty at various cell concentrations were cleaned up using buoyant density centrifugation and the Salmonella cell numbers were estimated with our method. Linear decreases in the threshold cycle value were observed during incubation of the samples. The standard curves for the cell number in all food samples were almost identical. With a standard curve using the mean parameter values, we successfully estimated viable Salmonella cell concentrations of the foods. The results indicate that our method is applicable for viable cell number estimation of the target microorganism in foods. Further, we used this method to study Salmonella growth in ground chicken stored at a constant temperature.

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RNA‐seq‐based selection of reference genes for RT‐qPCR analysis of pitaya

et al. 2019

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Reverse‐transcription quantitative real‐time PCR (RT‐qPCR) is a primary tool for measuring gene expression levels, and selection of appropriate reference genes is crucial for accurate and reproducible results of gene expression under various experimental conditions. However, no systematic evaluation of reference genes in pitaya (Hylocereus undatus Britt.) has been performed. Here, we examined the expression of five candidate reference genes, namely elongation factor 1‐alpha (HuEF1‐α), 18S ribosomal RNA (Hu18S rRNA), ubiquitin (HuUBQ), actin (HuACT), and ubiquitin‐conjugating enzyme (HuUQT), under different conditions in pitaya. The expression stabilities of these five genes were evaluated using two computation programs: geNorm and NormFinder. The results were further validated by normalizing the expression of the phosphoglycerate kinase (HuPGK) and ethylene‐responsive transcription factor (HuERF) genes. Our results indicate that combined use of HuUBQ and HuUQT is the most stable reference under all of the experimental conditions examined. HuEF1‐α, HuUBQ, and HuUQT are the top three most stable reference genes under salt stress, drought stress, and heat stress, and across different cultivars. HuEF1‐α, HuACT, and HuUQT exhibited the most stable expression patterns across different tissues. Our results will allow researchers to select the most appropriate reference genes for gene expression studies of pitaya under different conditions.

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Real-time Polymerase Chain Reaction for the Food Microbiologist: Technologies, Applications, and Limitations

Cited by 82 publications

References 35 publications

Origin Determination and Differentiation of Gelatin Species of Bovine, Porcine, and Piscine through Analytical Methods

Origin Determination and Differentiation of Gelatin Species of Bovine, Porcine, and Piscine through Analytical Methods

Estimation of Viable Salmonella Cell Numbers in Meat and Meat Product Using Real-Time PCR

RNA‐seq‐based selection of reference genes for RT‐qPCR analysis of pitaya

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