Real-Time PCR and Real-Time RT-PCR Applications in Food Labelling and Gene Expression Studies
International Journal of Genetics and Genomics
Volume 2, Issue 1, February 2014, Pages: 6-12
Received: Mar. 16, 2014;
Accepted: Apr. 14, 2014;
Published: Apr. 30, 2014
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Arash Kashani, Animal Science and Genetics, Tasmanian Institute of Agriculture, School of Land and Food, Faculty of Science, Engineering and Technology, University of Tasmania, Private Bag 54 Sandy Bay, Hobart, Tasmania 7001, Australia
Aduli Enoch Othniel Malau-Aduli, Animal Science and Genetics, Tasmanian Institute of Agriculture, School of Land and Food, Faculty of Science, Engineering and Technology, University of Tasmania, Private Bag 54 Sandy Bay, Hobart, Tasmania 7001, Australia; School of Veterinary and Biomedical Sciences, Faculty of Health, Medicine and Molecular Sciences, James Cook University, Townsville, Queensland 4811, Australia
Polymerase chain reaction (PCR) as a scientific invention, has revolutionized molecular biology and led to real-time PCR and later, real-time reverse transcription PCR (Real-Time RT-PCR). These two techniques enable scientists to conduct PCR detection of amplified gene products and expression analysis of targeted genes. Quantitative polymerase chain reaction (qPCR), also called real-time polymerase chain reaction, is a recent modification to PCR that utilizes fluorescent reporter molecular techniques to monitor the production of amplified products during each cycle of the PCR reaction, and enables both detection and quantification of specific sequences in complex mixtures. Over the past decade, real-time PCR applications have rapidly changed the nature of molecular science and become widely used tools in molecular genetics research. Real-time PCR permits specific, sensitive and reproducible manipulation of nucleic acids by combining the nucleic acid amplification and detection steps using gel electrophoresis. Hence, it almost eliminates the need for DNA sequencing or Southern blotting for amplicon identification. One of the many versions of PCR is real-time RT-PCR which has become one of the most broadly used gene amplification and expression methods in molecular biology research. Real-time RT-PCR is commonly employed to discover RNA expression levels through the creation of complimentary DNA (cDNA) transcripts from RNA, and it is frequently confused with real-time PCR. Food labelling provides very important information to help both producers and consumers to make informed choices about healthier and safer food. The process that information from a gene is used in the synthesis of a functional gene product is called gene expression. It enables scientists decipher the functions of genes. Food labelling and gene expression are fundamental to studying the relationships between the human genome, nutrition and health in a relatively new specialist field called nutritional genomics. Nutritional genomics is expected to revolutionize the way health professionals and dieticians treat people in the future. Thus, it is anticipated that the focus of nutritional genomics research will in the future, shift to determining the right type of food for an individual based on his or her genomic compatibility and therefore aid in avoiding foods that are an inappropriate match and could potentially impact negatively on the individual’s health. This paper reviews the importance and power of real-time PCR application in food labelling and nutritional genomics, types of fluorescent-based chemistry procedures developed for real-time PCR detection, real-time RT-PCR application in gene expression studies and the great potential of combining these technologies for animal molecular genetics research in sheep and fish
Aduli Enoch Othniel Malau-Aduli,
Real-Time PCR and Real-Time RT-PCR Applications in Food Labelling and Gene Expression Studies, International Journal of Genetics and Genomics.
Vol. 2, No. 1,
2014, pp. 6-12.
Deepak, S. A., Kottapalli, K. R., Rakwal, R., Oros, G., Rangappa, K. S., Iwahashi, H., Masuo, Y. & Agrawal, G. K. 2007., Real-Time PCR: Revolutionizing Detection and Expression Analysis of Genes. Current Genomics, 2007. 8(4): p. 234-251.
Fraga, D., T. Meulia, and S. Fenster, Real-Time PCR, in Current Protocols Essential Laboratory Techniques. 2008, John Wiley & Sons, Inc.
Mackay, I.M., K.E. Arden, and A. Nitsche, Real-time PCR in virology. Nucleic Acids Research, 2002. 30(6): p. 1292-1305.
Fajardo, V., González, I., Rojas, M., García, T. & Martín, R. 2010, A review of current PCR-based methodologies for the authentication of meats from game animal species. Trends in Food Science & Technology, 2010. 21(8): p. 408-421.
Hoffmann, B., Beer, M., Reid, S. M., Mertens, P., Oura, C. A. L., Vanrijn, P. A., Slomka, M. J., Banks, J., Brown, I. H., Alexander, D. J. & King, D. P. 2009., et al., A review of RT-PCR technologies used in veterinary virology and disease control: Sensitive and specific diagnosis of five livestock diseases notifiable to the World Organisation for Animal Health. Veterinary Microbiology, 2009. 139(1–2): p. 1-23.
Ponchel, F., Toomes, C., Bransfield, K., Leong, F., Douglas, S., Field, S., Bell, S., Combaret, V., Puisieux, A., Mighell, A., Robinson, P., Inglehearn, C., Isaacs, J. & Markham, A. 2003., Real-time PCR based on SYBR-Green I fluorescence: An alternative to the TaqMan assay for a relative quantification of gene rearrangements, gene amplifications and micro gene deletions. BMC Biotechnology, 2003. 3(1): p. 18.
Schefe, J., Lehmann, K., Buschmann, I., Unger, T. & Funke-kaiser, H. 2006., Quantitative real-time RT-PCR data analysis: current concepts and the novel “gene expression’s C T difference” formula. Journal of Molecular Medicine, 2006. 84(11): p. 901-910.
Shiao, Y.-H., A new reverse transcription-polymerase chain reaction method for accurate quantification. BMC Biotechnology, 2003. 3(1): p. 22.
Bustin, S. A., Benes, V., Nolan, T. & Pfaffl, M. W. 2005., Quantitative real-time RT-PCR – a perspective. Journal of Molecular Endocrinology, 2005. 34(3): p. 597-601.
K?ppel, R., J. Ruf, and J. Rentsch, Multiplex real-time PCR for the detection and quantification of DNA from beef, pork, horse and sheep. European Food Research and Technology, 2011. 232(1): p. 151-155.
Tanabe, S., Hase, M., Yano, T., Sato, M., Fujimura, T. & Akiyama, H. 2007., A Real-Time Quantitative PCR Detection Method for Pork, Chicken, Beef, Mutton, and Horseflesh in Foods. Bioscience, Biotechnology, and Biochemistry, 2007. 71(12): p. 3131-3135.
López-Calleja, I., González, I., Fajardo, V., Martín, I., Hernández, P. E., García, T. & Martín, R. 2007., Quantitative detection of goats’ milk in sheep’s milk by real-time PCR. Food Control, 2007. 18(11): p. 1466-1473.
Kesmen, Z., Yetiman, A. E., ?ahin, F. & Yetim, H. 2012., Detection of Chicken and Turkey Meat in Meat Mixtures by Using Real-Time PCR Assays. Journal of Food Science, 2012. 77(2): p. C167-C173.
Mutch, D.M., W. Wahli, and G. Williamson, Nutrigenomics and nutrigenetics: the emerging faces of nutrition. The FASEB Journal, 2005. 19(12): p. 1602-1616.
Woods, V.B. and A.M. Fearon, Dietary sources of unsaturated fatty acids for animals and their transfer into meat, milk and eggs: A review. Livestock Science, 2009. 126(1-3): p. 1-20.
Fenech, M., El-Sohemy, A., Cahill, L., Ferguson, L. R., French, T. A. C., Tai, E. S., Milner, J., Koh, W. P., XIE, L., Zucker, M., Buckley, M., Cosgrove, L., Lockett, T., Fung, K. Y. C. & Head, R. 2011., Nutrigenetics and Nutrigenomics: Viewpoints on the Current Status and Applications in Nutrition Research and Practice. Journal of Nutrigenetics and Nutrigenomics, 2011. 4(2): p. 69-89.
Wong, M.L. and J.F. Medrano, Real-time PCR for mRNA quantitation. Biotechniques, 2005. 39(1): p. 75-85.
Etienne, W., Meyer, M. H., Peppers, J. & Meyer, R. A. 2004., Comparison of mRNA gene expression by RT-PCR and DNA microarray. Biotechniques, 2004. 36(4): p. 618-20, 622, 624-6.
Alhazzaa, R., Bridle, A. R., Nichols, P. D. & Carter, C. G. 2011., et al., Up-regulated Desaturase and Elongase Gene Expression Promoted Accumulation of Polyunsaturated Fatty Acid (PUFA) but Not Long-Chain PUFA in Lates calcarifer, a Tropical Euryhaline Fish, Fed a Stearidonic Acid- and γ-Linoleic Acid-Enriched Diet. Journal of Agricultural and Food Chemistry, 2011. 59(15): p. 8423-8434.
Codabaccus, B. M., Bridle, A. R., Nichols, P. D. & Carter, C. G. 2011., et al., An extended feeding history with a stearidonic acid enriched diet from parr to smolt increases n-3 long-chain polyunsaturated fatty acids biosynthesis in white muscle and liver of Atlantic salmon (Salmo salar L.). Aquaculture, 2011. 322–323(0): p. 65-73.
Ballent, M., Wilkens, M. R., Maté, L., Muscher, A. S., Virkel, G., Sallovitz, J., Schr?der, B., Lanusse, C. & Lifschitz, A. 2013., P-glycoprotein in sheep liver and small intestine: gene expression and transport efflux activity. Journal of Veterinary Pharmacology and Therapeutics, 2013: p. n/a-n/a.
He, Y., Liu, D., Xi, D., Yang, L., Tan, Y., Liu, Q., Mao, H. & Deng, W. 2010., Isolation, sequence identification and expression profile of three novel genes Rab2A, Rab3A and Rab7A from black-boned sheep (Ovis aries). Molecular Biology, 2010. 44(1): p. 14-22.
Yu, H., Chen, S., Xi, D., He, Y., Liu, Q., Mao, H. & Deng, W. 2010., et al., Molecular cloning, sequence characterization and tissue transcription profile analyses of two novel genes: LCK and CDK2 ; from the Black-boned sheep (Ovis ariest). Molecular Biology Reports, 2010. 37(1): p. 39-45.
Xi, D., He, Y., Sun, Y., Gou, X., Yang, S., Mao, H. & Deng, W. 2011., et al., Molecular cloning, sequence identification and tissue expression profile of three novel genes Sfxn1, Snai2 and Cno from Black-boned sheep (Ovis aries). Molecular Biology Reports, 2011. 38(3): p. 1883-1887.
Cao, G. L., Zhang, Y. J., Wang, J. M. & Jiang, Y. L. 2010., et al., Analysis on cDNA sequence, mRNA expression and imprinting status of Dlk1 gene in goats. Molecular Biology Reports, 2010. 37(5): p. 2259-2264.
Han, C.-X., H.-X. Liu, and D.-M. Zhao, The quantification of prion gene expression in sheep using real-time RT-PCR. Virus Genes, 2006. 33(3): p. 359-364.