In-house polymerase chain reaction (PCR) assays are now an integral part of the work of most diagnostic microbiological laboratories. Despite the availability of commercial reagent 'master-mixes' of some PCR reagents, the optimisation of primers still poses a significant problem. Here, we describe a simple method to assess the concentration of primer needed in single round, multiplex, nested an...

We have developed a highly sensitive method for DNA analysis on 3D gel element microarrays, a technique we call multiplex microarray-enhanced PCR (MME-PCR). Two amplification strategies are carried out simultaneously in the reaction chamber: on or within gel elements, and in bulk solution over the gel element array. MME-PCR is initiated by multiple complex primers containing gene-specific, forw...

PCR-based strategies such as competitive RT-PCR or semiquantitative RTPCR are commonly used for quantitating low-abundance mRNA transcripts (4,7–9). Competitive RT-PCR relies on amplifying the same target with a known amount of a competitor/reference sequence and comparing the relative amounts of the two PCR products. To improve the sensitivity and accuracy of these methods, often expensive and...

Copying multiple regions of a DNA molecule is routinely performed today using the polymerase chain reaction (PCR) in a process commonly referred to as multiplex PCR. The development of a multiplex PCR reaction involves designing primer sets and examining various combinations of those primer sets and different reaction components and/or thermal cycling conditions. The process of optimizing a mul...

We developed a primer design method, Pythia, in which state of the art DNA binding affinity computations are directly integrated into the primer design process. We use chemical reaction equilibrium analysis to integrate multiple binding energy calculations into a conservative measure of polymerase chain reaction (PCR) efficiency, and a precomputed index on genomic sequences to evaluate primer s...

UNLABELLED Designed degenerate primers unlike conventional primers are superior in matching and amplification of large number of genes, from related gene families. DPPrimer tool was designed to predict primers for PCR amplification of homologous gene from related or diverse plant species. The key features of this tool include platform independence and user friendliness in primer design. Embedde...

TaqMan, a variation of fluorescent PCR, is a powerful tool for gene expression and polymorphism studies. Here we describe the design and evaluation of 27 new TaqMan primer-probe sets for rat genes that play a key role in neural signaling. These newly designed and synthesized probes were tested and then used for quantification of RNA isolated from rat brain. The usual length of common TaqMan pro...

The success of the polymerase chain reaction (PCR) is highly dependent on primer design. Commonly used primer design programs rely upon a core set of parameters such as melting temperature, primer length, GC content, and complementarity to optimize the PCR product, but weight those parameters to differing degrees, as well as include other parameters for PCR specific tasks. An analysis of these ...

The polymerase chain reaction (PCR) is an indispensable DNA amplification technique that has been exploited in numerous areas, including molecular diagnostics. One major drawback of PCR is the competing amplification of undesired off-target products, which primarily occurs at ambient temperatures prior to PCR cycling (1). Hot Start PCR techniques aim to block the extension of primers in nonspec...

Background: It is estimated that about 3,000 pregnancies in Iran are at risk for b-thalassemia each year. Objective: To evaluate the diagnostic accuracy of combination of ARMS/PCR and  RFLP/PCR techniques in prenatal diagnosis of b-thalassemia.Methods: Sixty-seven b-thalassemia carrier families were enrolled in this study. To analyze b-globin gene, amplification refractory mutation system (ARMS...