Real-time PCR, or quantitative PCR (QPCR), has been developed to rapidly detect and quantify the total number of yeasts in wine without culturing. Universal yeast primers were designed from the variable D1/D2 domains of the 26S rRNA gene. These primers showed good specificity with all the wine yeasts tested, and they did not amplify the most representative wine species of acetic acid bacteria a...

Real-time quantitative polymerase chain reaction (qPCR) testing for Mycobacterium avium subspecies paratuberculosis (MAP) in fecal samples is a rapid alternative to culture on Herrold egg yolk medium (HEYM), the traditional antemortem reference test for MAP. Although the sensitivity and specificity of these 2 tests have been estimated based on dichotomized test results, the correlation between ...

Real-time, or quantitative, polymerase chain reaction (qPCR) is quickly supplanting other molecular methods for detecting the nucleic acids of human and other animal pathogens owing to the speed and robustness of the technology. As the aquatic animal health community moves toward implementing national diagnostic testing schemes, it will need to evaluate how qPCR technology should be employed. T...

Identification of appropriate reference genes (RGs) is critical to accurate data interpretation in quantitative real-time PCR (qPCR) experiments. In this study, we have utilised next generation RNA sequencing (RNA-seq) to analyse the transcriptome of a panel of non-melanoma skin cancer lesions, identifying genes that are consistently expressed across all samples. Genes encoding ribosomal protei...

Quantitative Real-Time Polymerase Chain Reaction, better known as qPCR, is the most sensitive and specific technique we have for the detection of nucleic acids. Even though it has been around for more than 30 years and is preferred in research applications, it has yet to win broad acceptance in routine practice. This requires a means to unambiguously assess the performance of specific qPCR anal...

We review different methodologies to estimate the expression levels of microRNAs (miRNAs) using real-time quantitative PCR (qPCR). As miRNA analysis is a fast changing research field, we have introduced novel technological approaches and compared them to existing qPCR profiling methodologies. qPCR also remains the method of choice for validating results obtained from whole-genome screening (e.g...

A novel SYBR® green-real time polymerase chain reaction (qPCR) was developed to detect two Bartonella species, B. henselae and B. clarridgeiae, directly from blood samples. The test was used in blood samples obtained from cats living in animal shelters in Southern Brazil. Results were compared with those obtained by conventional PCR targeting Bartonella spp. Among the 47 samples analyzed, eight...

An efficient protocol for the manual fluorescent MGIT culture system combined with rapid confirmation of Mycobacterium avium subsp. paratuberculosis (MAP) growth in the broth culture was established and evaluated for the detection of viable MAP in direct quantitative PCR (QPCR) positive bovine feces. Manually detected fluorescence emissions from MGIT tubes were analyzed objectively using an ope...

A quantitative real-time PCR (qPCR) based on a standard curve was developed for detection and quantitation of ranaviruses. The target gene for the qPCR was viral DNA polymerase (DNApol). All ten ranavirus isolates studied (Epizootic haematopoietic necrosis virus, EHNV; European catfish virus, ECV; European sheatfish virus, ESV; Frog virus 3, FV3; Bohle iridovirus, BIV; Doctor fish virus, DFV; G...

BACKGROUND Quantitative PCR (qPCR) is a workhorse laboratory technique for measuring the concentration of a target DNA sequence with high accuracy over a wide dynamic range. The gold standard method for estimating DNA concentrations via qPCR is quantification cycle () standard curve quantification, which requires the time- and labor-intensive construction of a standard curve. In theory, the sha...