Quantitative relative and absolute real-time PCR (qRT-PCR)IntroductionQuantitative real-time PCR (qRT-PCR)Polymerase chain reaction (PCR) is a technology revolutionized used in molecular biology for the detection and amplification of DNA generating thousands or even millions of copies of a particular DNA sequence. Real-time reverse transcription-polymerase chain reaction quantitative PCR (RT-QPCR) is considered the gold standard technique in molecular biology and has been considered a gold standard assay for the detection of DNA and mRNA. This technique is also used in a wide range of fields. variety of bioanalytical scientific fields (Burns et al, 2005). RT-QPCR is a technique that simultaneously monitors, amplifies, and quantifies nucleic acid accumulation in real time and is characterized as a high-quality, high-performance technique associated with high throughput, reproducibility, specificity, and sensitivity. qRT-PCR involves a number of steps, including RNA extraction, cDNA synthesis, PCR amplification, and normalization to an appropriate internal standard or reference gene. Different screening methods and internal standards all have an effect on the reliability of qRT-PCR (Zhang et al, 2012). RT-qPCR validates and analyzes data in basic fluorescent thermal cycling PCR software that depends on absolute or relative quantification of PCR data. This validation system requires comparison of the sample to standard DNA or reference genes, respectively (Roussel et al, 2007). .Relative quantification real-time PCR (qRT-PCR)Relative quantification has been widely studied in cytokines or enzymes to assess changes in gene expression; this type of quantification omits the use of standards and allows the comparison of two experiments that...... middle of article ......e RT-PCR quantification of Kuruma shrimp transcripts: A comparison of relative and absolute quantification procedures'. Journal of Biotechnology, 129 (3), pp.391-399. Smith, CJ & Osborn, AM (2009) “Advantages and limitations of quantitative PCR (Q-PCR)-based approaches in microbial ecology”. FEMS Microbiology Ecology, 67 (1), pp.6-20. Valasek, MA & Repa, JJ (2005) “The power of real-time PCR”. Advances in Physiology Education, 29 (3), pp.151-159. Wong, ML & Medrano, JF (2005) Real-time PCR for mRNA quantification. BioTechniques, 39, 75-85Zhang, J., Hu, Y.-h., Sun, B.-g., Xiao, Z.-z. & Sun, L. (2013) “Selection of normalization factors for quantitative real-time RT-PCR studies in Japanese flounder (Paralichthys olivaceus) and turbot (Scophthalmus maximus) under viral infection conditions”. Veterinary immunology and immunopathology, 152 (3–4), pp..303-316