PCR – Multimedia

Multiplex PCR is an advantageous technique used in PCR applications to amplify multiple targets in a single reaction. As useful as it is, this technique presents a new set of challenges that further complicates PCR setup. For example, reactions are more prone to off-target amplifications such as mis-priming and primer dimer due to the increased number of primer pairs.

PCR is a widely used scientific tool whose specificity can be increased by the use of Hot Start technologies. Although many Hot Start technologies exist, recently developed CleanAmp™ dNTPs are a distinct approach that employs modified nucleoside triphosphates with a thermolabile protecting group at the 3´-hydroxyl.

Improved FFPE RNA isolation, novel TR priming for complete recovery of FFPE mRNA fragments. Differential expression defines Scores for tumor classification: comparing fresh-frozen with FFPE. Novel flow-through Ziplex microarray platform demonstrates MAQC performance level of data obtained with FFPE samples.

Multiple Sclerosis (MS) is a chronic inflammatory demyelinating disorder of the CNS. No successful treatment for MS, but one therapeutic strategy in research is the use of bone marrow-derived mesenchymal stem cells (MSC). We studied a group of MS patients who underwented MSCs, assayed for expression of a transcription factor, FOXP3, as a specific marker of MS amelioration in peripheral blood. qRT-PCR on PBMCs showed higher FoxP3 levels.

PCR is a widely used scientific tool employed by a variety of applications. Various Hot Start technologies have already been developed using modified PCR components to increase specificity of a reaction. Recently developed CleanAmpTM dNTPs are modified nucleoside triphosphates with a thermolabile 3’-tetrahydrofuranyl protecting group that is released at higher temperatures. These modified dNTPs prevent low temperature primer extension, which can often be a significant problem in PCR. At higher t

Multiplex PCR is an advantageous technique used in PCR applications to amplify multiple targets in a single reaction. As useful as it is, this technique presents a new set of challenges that further complicates PCR setup. For example, reactions are more prone to off-target amplifications such as mis-priming and primer dimer due to the increased number of primer pairs. Furthermore, preferential amplification of certain targets leads to an unequal distribution of amplicon products, making quantifi

dPCR is achieved by sample partitioning prior to PCR amplification such that each reaction chamber contains one copy or less of target DNA. This dilution becomes the limiting factor and an accurate target molecule count is achievable. This study evaluates dPCR’s quantitative capabilities and investigates parameters influencing copy number quantification, using the Fluidigm Biomark instrument. Biomark technology combines dPCR theory with a microfluidics platform.

Our project is partially focused on insulin affecting four proteins associated with lung surfactant. Small and hydrophobic SP-B and S-PC help to spread and stabilize the surfactant layer while large and hydrophilic SP-A and SP-D participate in immune responses. A549 and H441 cell lines were used. It was observed using RealTime PCR that higher doses of insulin lead to decreased transcription of SP-B, SP-C, SP-D, and both isoforms of SP-A.

A triplex qPCR assay was developed to screen for three major food-borne pathogens in fish. The assay is based on simultaneous qPCR amplification of specific target genomes using fluorophore labelled locked nucleic acid (LNA) TaqMan probes. The detection limit of the qPCR is 1 colony forming unit (cfu) per 1ml and the detection limit of the assay with 24h general enrichment is 1 cfu per 25g of fish meat.

Sequenom EpiTYPER analysis of GNAS methylation in Pseudohypoparathyroidism type Ib patients reveals overall GNAS methylation defects also for the familial cases. Such abnormalities are not detectable via old methodologies such as PCR followed by methylation specific restriction digestion and are here for the first time described.