Journal ArticleDOI
Gene quantification using real-time quantitative PCR: an emerging technology hits the mainstream.
TLDR
The recent flood of reports using real-time Q-PCR testifies to the transformation of this technology from an experimental tool into the scientific mainstream and this review will help guide the reader through the variables that can limit the usefulness of thistechnology.About:
This article is published in Experimental Hematology.The article was published on 2002-06-01. It has received 1178 citations till now.read more
Citations
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Quantification of mRNA using real-time RT-PCR
TL;DR: A series of RT-qPCR protocols are described that illustrate the essential technical steps required to generate quantitative data that are reliable and reproducible in molecular medicine, biotechnology, microbiology and diagnostics.
Journal ArticleDOI
Guideline to reference gene selection for quantitative real-time PCR.
Aleksandar Radonic,Stefanie Thulke,Ian M. Mackay,Olfert Landt,Wolfgang Siegert,Andreas Nitsche,Andreas Nitsche +6 more
TL;DR: In this paper, quantitative real-time PCR was used to determine the mRNA transcription profiles of 13 putative reference genes, comparing their transcription in 16 different tissues and in CCRF-HSB-2 cells stimulated with 12-Otetradecanoylphorbol-13-acetate and ionomycin.
Journal ArticleDOI
Quantitative real-time RT-PCR – a perspective
TL;DR: The real-time reverse transcription polymerase chain reaction uses fluorescent reporter molecules to monitor the production of amplification products during each cycle of the PCR reaction, which combines the nucleic acid amplification and detection steps into one homogeneous assay and obviates the need for gel electrophoresis to detect amplification products.
Journal ArticleDOI
Comprehensive Algorithm for Quantitative Real-Time Polymerase Chain Reaction
Sheng Zhao,Russell D. Fernald +1 more
TL;DR: This algorithm provides an objective and noise-resistant method for quantification of qRT-PCR results that is independent of the specific equipment used to perform PCR reactions.
Journal Article
Pitfalls of quantitative real-time reverse-transcription polymerase chain reaction.
Stephen A. Bustin,Tania Nolan +1 more
TL;DR: Real-time RT-PCR remains a research tool, and it is important to recognize the considerable pitfalls associated with transcriptome analysis, with the successful application of RTPCR depending on careful experimental design, application, and validation.
References
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Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction
TL;DR: A new method of total RNA isolation by a single extraction with an acid guanidinium thiocyanate-phenol-chloroform mixture is described, providing a pure preparation of undegraded RNA in high yield and can be completed within 4 h.
Journal ArticleDOI
Real time quantitative PCR.
TL;DR: Unlike other quantitative PCR methods, real-time PCR does not require post-PCR sample handling, preventing potential PCR product carry-over contamination and resulting in much faster and higher throughput assays.
Journal ArticleDOI
Molecular Beacons: Probes that Fluoresce upon Hybridization
Sanjay Tyagi,Fred Russell Kramer +1 more
TL;DR: Novel nucleic acid probes that recognize and report the presence of specific nucleic acids in homogeneous solutions that undergo a spontaneous conforma-tional change when they hybridize to their targets are developed.
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Absolute quantification of mRNA using real-time reverse transcription polymerase chain reaction assays.
TL;DR: The technical aspects involved are discussed, conventional and kinetic RT-PCR methods for quantitating gene expression are contrasted, and the usefulness of these assays are illustrated by demonstrating the significantly different levels of transcription between individuals of the housekeeping gene family, glyceraldehyde-3-phosphate-dehydrogenase (GAPDH).
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Kinetic PCR Analysis: Real-time Monitoring of DNA Amplification Reactions
TL;DR: Results obtained with this approach indicate that a kinetic approach to PCR analysis can quantitate DNA sensitively, selectively and over a large dynamic range.