Quantitative real-time PCR represents a highly sensitive and powerful technique for the quantitation of nucleic acids. It has a tremendous potential for the high-throughput analysis of gene expression in research and routine diagnostics. However, the major hurdle is not the practical performance of the experiments themselves but rather the efficient evaluation and the mathematical and statistic...

The real-time quantitative polymerase chain reaction (rtqPCR) has overcome the limitations of conventional, time-consuming quantitative PCR strategies and is maturing into a routine tool to quantify gene expression levels, following reverse transcription (RT) of mRNA into complementary DNA (cDNA). Expression profiling with single-cell resolution is highly desirable, in particular for complex ti...

OBJECTIVES To develop a real-time quantitative RT-PCR method for BRCA1 mRNA and then use it for the study of BRCA1 gene expression in human MCF-7 breast cancer cells after their exposure to antineoplastic agents and gamma irradiation. DESIGN AND METHODS The developed QRT-PCR method is based on the real-time monitoring of a fluorescein-labeled TaqMan probe, specific for BRCA1 mRNA, during PCR ...

BACKGROUND Biological markers capable of predicting the risk of recurrence and the response to treatment in breast cancer are eagerly awaited. Estrogen and progesterone receptors (ER, PgR) in tumor cells mark cancers that are more likely to respond to endocrine treatment, but up to 40% of such patients do not respond. Here, the expression of a group of estrogen-regulated genes, previously ident...

In order to facilitate optimal plant DNA quantitation and identification, an assay has been developed that uses generic plant PCR primers that amplify a region in the chloroplast genome of plant samples. The assay uses the SYBR green detection dye to detect the PCR product with a universal PCR primer set to the large subunit of ribulose bisphosphate carboxylase, rbcL, but can be used with any o...

Standard screening of melanoma patients is a useful tool for predicting outcome of patients, however, an instant methodology for exact detection of subclinical disease or monitoring treatment response is under investigation. Detection of circulating melanoma cells is, therefore, a possible novel promising staging method. However, inconsistent data on method sensitivity and on the predicted pati...

8 Improved understanding of the biology of reproduction in filarial worms may lead to identification of new targets for drugs or vaccines. Real-time RT-PCR is increasingly being adopted for RNA quantification and genetic analysis. Candidate gender-regulated genes were selected from genes identified in prior studies by differential display RT-PCR and by electronic selection of the Brugia malayi ...

Objective. Low gene expression in rare cell subpopulations can make it difficult to identify transcripts using real time quantitative RT-PCR (qRT-PCR). Transcript number can be increased using linear amplification, but this technique amplifies the 3’ end of mRNA, imposing severe limitations on qRT-PCR primer design. Artifacts such as primer-dimers, introduced by these limitations, may interfere...

Circulating tumor cells (CTCs) were recognized as novel tumor biomarker for prognostic and predictive purposes in various cancers. Various detection technologies and devices have been developed to enumerate and characterize CTCs. Most of those approaches are based on the positive enrichment strategy and immunocytological techniques. However, the sensitivity of these approaches proved to be limi...

The stability of standard gene expression is an elementary prerequisite for internal standardisation of target geneexpression data and many so called housekeeping genes with assumed stable expression can exhibit either up-or down-regulation under some experimental conditions. The developed, and herein presented, software calledBestKeeper determines the best suited standards, out...