Gut colonization with enterotoxigenic Bacteroides fragilis (ETBF) appears to be associated with the development of colorectal cancer. However, differences in carriage rates are seen with various testing methods and sampling sites. We compared standard PCR, SYBR green and TaqMan quantitative PCR (qPCR) and digital PCR (dPCR) in detecting the B. fragilis toxin (bft) gene from cultured ETBF, and f...

Background: The outbreak of novel influenza A H1N1-2009 virus (pH1N1) and its rapid spread worldwide raised serious concern about pandemic preparedness. Objectives: The present study was designed to evaluate the sensitivity and specificity of two chemistries of real-time RT-PCR (with the use of fluorescent SYBR Green I dye and specific TaqMan probe) for detection of the matrix and hemagglutinin...

We have previously described the development of a onetube SYBR Green real-time RT-PCR assay for the detection and quantitation of infectious salmon anemia virus (ISAV) in various biological samples. The twofold aim of the present study was to verify that the optimized SYBR Green real-time RT-PCR conditions could detect ISAV isolates of different geographic origins, and to analyze the growth pat...

Real-time PCR is an accurate method that can be used for the quantification of specific DNA molecules. Here we provide a protocol for SYBR Green I in real-time PCR applications using plastic reaction tubes. We report that SYBR Green I is alkali labile and once degraded inhibits the PCR. In our optimized protocol, diluted aliquots of SYBR Green I remain stable for at least two weeks. We also eva...

The precise diagnosis of American tegumentary leishmaniasis (ATL) is an essential task due to the disease's associated morbidity. A noninvasive, extremely sensitive, and highly specific exam is critical, particularly for mucosal leishmaniasis (ML), in which a low parasite quantity is expected. We aimed to compare the diagnostic accuracy of swab and biopsy sample analysis using SYBR Green- and T...

A new membrane-permeant DNA stain, SYBR-14, was used in combination with propidium iodide (PI) to estimate the proportion of living sperm in bovine semen. The SYBR-14 stained living sperm while PI only stained degenerate cells that had lost their membrane integrity. Staining with SYBR-14 resulted in the nuclei of living sperm fluorescing bright green. Aliquots containing nearly all living bovin...

Diagnostic tests based on detection of dengue virus (DENV) genome are available with varying sensitivities and specificities. The Simplexa Dengue assay (Focus Diagnostics) is a newly developed real-time RT-PCR method designed to detect and serotype DENV simultaneously. To assess the performance of the Simplexa Dengue assay, we performed comparison with conventional RT-PCR and SYBR Green real-ti...

HHV-6 is the etiological agent of Exanthem subitum which is considered the sixth most frequent disease in infancy. In immuno-compromised hosts, reactivation of latent HHV-6 infection may cause severe acute disease. We developed a Sybr Green Real Time PCR for HHV-6 and compared the results with nested conventional PCR. A 214 pb PCR derived fragment was cloned using pGEM-T easy from Promega syste...

Four quantitative reverse transcription-PCR (RT-PCR) methods were compared to evaluate the time course of mRNA formation and decay. Mouse fibroblasts (NIH 3T3) transfected with the human beta-globin open reading frame/c-myc 3'-untranslated region chimeric gene under control of the c-fos promoter (fos-glo-myc) were used for serum-inducible transcription. The amount of fos-glo-myc mRNA, relative ...

Lyme disease caused by Borrelia burgdorferi is the most common tick-borne disease in the US and Europe. Unlike most bacteria, measurements of growth and viability of B. burgdorferi are challenging. The current B. burgdorferi viability assays based on microscopic counting and PCR are cumbersome and tedious and cannot be used in a high throughput format. Here, we evaluated several commonly used v...