OBJECTIVES Determination of bacterial antimicrobial susceptibility is usually performed using phenotypic methods. In this study, we developed a universal 16S rRNA and rpoB quantitative PCR assay for susceptibility testing of bacteria commonly isolated in clinical microbiology laboratories. METHODS Antibiotic susceptibilities for 24 bacterial strains of various species were tested by real-time...

BACKGROUND Research into gene expression enables scientists to decipher the complex regulatory networks that control fundamental biological processes. Quantitative real-time PCR (qPCR) is a powerful and ubiquitous method for interrogation of gene expression. Accurate quantification is essential for correct interpretation of qPCR data. However, conventional relative and absolute quantification m...

Lentiviral vectors are efficient vehicles for stable gene transfer in both dividing and non-dividing cells. This feature among others makes lentiviral vectors a powerful tool in molecular research. However, the use of lentiviruses in research studies and clinical trials requires a precise and validated titration method. In this study, we describe a qPCR-based approach for estimation of lentivir...

as a template for PCR. Sequencing of the plasmid DNA can now be performed using two PCR primers flanking the MCS of the vector (we used M13 universal forward primer and M13 universal reverse primer; Figure 1). If the unique primer is set 100–150 bp from the end of the known sequence, sufficient overlapping sequence is generated to align the sequences unambiguously. This method was used in our l...

Three tuberculous, twenty-one non-tuberculous mycobacterial (NTM) reference strains and seventy two isolates classified by biochemical tests were shown to produce specific sets of DNA fragments in a polymerase chain reaction with single universal primer (UP-PCR). A rather wide limit of tolerance for variations in procedure of PCR mixture preparation and thermocycling parameters was found. There...

Multiplex PCR amplification followed by either agarose gel electrophoresis (PCR-AGE) or microchip electrophoresis (PCR-ME) was used to test a total of 120 fungal strains. The internal transcribed spacer 1 (ITS1) and ITS2 regions and the 5.8S ribosomal DNA (rDNA) region of the fungi were amplified by using universal primers ITS1 and ITS4. The ITS2 region was simultaneously amplified by using uni...

A new real-time PCR detection system was developed for grapevine yellows (GY) using TaqMan minor groove binder probes and including two amplicons for group-specific detection of Flavescence dorée (FD) and Bois noir (BN) phytoplasmas, plus a universal phytoplasma amplicon. FD and BN amplicons were designed to amplify species-specific genomic DNA fragments and the universal amplicon to amplify th...

Our understanding of thermophile diversity is based predominantly on PCR studies of community DNA. "Universal" and domain-specific rRNA gene PCR primers have historically been used for the assessment of microbial diversity without adequate regard to the degree of specificity of primer pairs to different prokaryotic groups. In a reassessment of the published primers commonly used for "universal"...