PCR - Articles and news items

Expression profiling of circulating miRNAs as a novel non-invasive diagnostic tool

Latest issue / 13 December 2011 / Mirco Castoldi. Department of Pediatric Hematology, Oncology and Immunology University of Heidelberg

Cell-free nucleic acids circulating in human blood were first described in 19481. However, it was not until the work of Sorengon and colleagues was published in 19942 that the importance of circulating nucleic acid (cfNA) was recognised. Today, the detection of diverse type of cfNA3 in blood and other body fluids is a valuable resource for the identification of a novel biomarker4,5. Although different types of cfNA have been described (including DNA, mRNA and microRNA), this review focuses on the isolation, detection and clinical utility of circulating microRNAs.

microRNAs (miRNAs) are an abundant class of short single stranded non-coding RNAs (~22 nts) that regulate gene expression at the posttranscriptional level. Interaction between an miRNA and any given of its mRNA targets results in either translation inhibition, mRNA degradation or a combination of both mechanisms. Therefore, miRNAs activity effectively reduces the transcriptional output of a target gene, without affecting its transcription rate. Currently, the sequence of over 60,000 microRNAs are deposited in the miRBase database [Version 17, April 20116]. miRNA activity has been associated with the control of a wide range of basic processes such as development, differentiation and metabolism. Detection of differential expression of miRNAs in many cases have established the basis for miRNA functional analysis and specific miRNA expression patterns can provide valuable diagnostic and prognostic indications, for example, in the context of human malignancies7,8. Moreover, the deregulation of the expression of miRNAs has been shown to contribute to cancer development through various kinds of mechanisms, including deletions, amplification or mutations involving miRNA loci, epigenetic silencing, as well as the dysregulation of transcription factors that target specific miRNAs9,10. (more…)

Abbott Receives FDA Approval for Molecular Test for Hepatitis C

Industry news, News / 18 May 2011 / Abbott

Abbott announced today that it has received approval from the U.S. Food and Drug Administration to market its RealTime PCR (polymerase chain reaction) test for measuring the viral load of hepatitis C (HCV), the leading cause of liver cancer in the United States.

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PCR and personalised cancer medicine

Issue 6 2010 / 16 December 2010 / Frank McCaughan, MRC Career Development Fellow, MRC Laboratory of Molecular Biology

The delivery of personalised medicine is a key goal of modern cancer medicine and refers to the tailoring of anticancer therapy to the molecular characteristics of an individual tumour. To facilitate personalised medicine, it is important to have robust and reproducible means of gaining molecular information about a patient’s cancer that can be used to guide clinical decision-making. There have therefore been tremendous efforts to identify molecular signatures – biomarkers – that can be used to help predict a cancer patient’s prognosis or their likelihood of a response to targeted drug therapies. Such molecular profiling has long been applied to haematological malignancies and is increasingly becoming the norm in the most common epithelial cancers such as lung and colorectal cancer. This article will focus on the role of the polymerase chain reaction (PCR) in helping to meet the challenges involved in the design, testing and delivery of personalised cancer medicine. (more…)

High-throughput PCR based diagnostics: Linking sample handling to molecular oncology risk groups

Issue 3 2010, Past issues / 24 June 2010 / Ehsan G. Karimiani, Stephan Mohr & Philip J. R. Day, University of Manchester

Cancer molecular pathology broadly relies on the comparison between diseased and normal tissues, with statistically validated differences revealing cancerassociated pathways. This approach, although comparatively one-dimensional, has been remarkably successful, enabling identification of many types of malignant biomarkers and providing the means to develop pharmaceutical agents directed against pertinent biological targets. Most typically during the progression of malignancies, pathologists employ morphological screening of cancerous tissues. However, this form of monitoring has significant limitations, particularly in the early stages of pre-treatment or during the clinical remission. (more…)

There is more to qPCR than the PCR reaction

Issue 5 2008, Past issues / 29 September 2008 /

The polymerase chain reaction is arguably the most significant technical discovery yet to have been made in the field of molecular biology and genetics, if not all life science. It cannot be overstated how much of an impact this technique has had, resulting in molecular biology becoming an integral part of most biological and medical research fields. PCR is both highly versatile and extremely sensitive, yet with basic training and adherence to key precautions against contamination, it is comparatively simple to perform. However, when conducting PCR, there are a number of additional considerations that are frequently overlooked when investigating the presence or amount of nucleic acid within a sample.

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Real-time PCR gene expression profiling

Issue 1 2007, Past issues / 25 January 2007 / Mikael Kubista, TATAA Biocenter and MultiD Analyses AB, Sweden, Björn Sjögreen, Center for Applied Scientific Computing, Lawrence Livermore National Laboratory, United States and MultiD Analyses AB, Amin Forootan, MultiD Analyses AB, Radek Sindelka and Jiri Jonák, Laboratory of Gene Expression, Institute of Molecular Genetics, Academy of Sciences of the Czech Republic and José Manuel Andrade, Dept of Analytical Chemistry, University of A Coruna, Spain

Real-time PCR has rapidly become the preferred technique for quantitative analysis of nucleic acids. Its superior sensitivity, reproducibility and dynamic range make it the preferred choice for expression profiling in scientific, as well as routine, applications.1

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Quantitative PCR assays in clinical drug development

Issue 5 2006, Past issues / 28 September 2006 / Michael E. Burczynski, Ph.D. and Ole E. Vesterqvist, Ph.D. Biomarker Laboratory, Clinical Translational Medicine, Wyeth Research

Biomarkers (biological markers) have become an integral part of both drug discovery and drug development and play an important role in the transition of potential new drugs from discovery into clinical drug development. In the past, most biomarkers were proteins/peptides and metabolites measured by technologies such as immunoassays, enzymatic assays, HPLC and mass spectrometry.

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Real-time PCR: clinical applications

Issue 2 2006, Past issues / 24 March 2006 / Lut Overbergh and Chantal Mathieu, Laboratory for Experimental Medicine and Endocrinology (LEGENDO), University Hospital Gasthuisberg, Catholic University of Leuven

The real-time PCR technique is one of the emerging techniques that, although only described for the first time about a decade ago, have become the method of choice for quantification of DNA and RNA levels in cells, tissues and tissue biopsies.

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Making the lab workhorse run faster

Issue 1 2006, Past issues / 2 February 2006 / Holly Hilton, Ph.D. and Charu Kanwal Ph.D. Research Informatics, Genetics and Genomics Hoffmann La-Roche Inc.

Quantitative real-time PCR, the workhorse of any genomics lab, is a well established technique that has numerous uses due to its simplicity and flexibility. In this article we will review a brief history of real time PCR, discuss our strategy for optimising a lab running Quantitative-RT-PCR and describe how this technique fits into the drug discovery process. We will conclude with some on-going issues regarding RT-PCR data generation and analysis.

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Options for quantitative analysis by real-time PCR

Issue 4 2005, Past issues / 11 November 2005 / Gareth Elvidge, PhD, Genomics Group, Wellcome Trust Centre for Human Genetics, University of Oxford

The expansion of microarray-based gene expression studies has led to an increase in demand for gene-specific PCR-based methods for independent validation of results. Although a number of technologies are available to meet this requirement the most popular is currently real-time PCR.

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