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[Real time quantitative PCR].

Authors: D W, Kim;

[Real time quantitative PCR].

Abstract

So far, quantitative techniques, such as PCR and FISH, have been used to detect of DNA and RNA. However, it is difficult to measure and compare the exact amount of amplified products with the results of endpoint analysis in conventional PCR techniques. Theoretically, there is a quantitative relationship between amount of starting target sequence and amount of PCR product at any given cycle. The development of real-time quantitative PCR (RQ-PCR) has eliminated the variability associated with conventional quantitative PCR, thus allowing the routine and reliable quantitation of PCR products. Detection of fluorescence during the thermal cycling process can be performed using iCycler(Bio-Rad), the GeneAmp 5700 or 7700(ABI-PRISM), and Light-Cycler(Roche). Two fluorogenic probes are available for use on real time quantitation. The fluorogenic 5'-nuclease assay(Taqman method) uses a fluorogenic probe to enable the detection of a sequence specific PCR product. Fluorogenic probe is incorporated with the reporter dye on the 5' end and the quencher on the 3' end. The second method uses SYBR Green I dye which is a highly specific double-stranded DNA binding dye. Real-time PCR is able to be possible exact quantitation of DNA and RNA much more precise and reproducible because it is based on CT values acquired during the exponential phase of PCR rather than endpoint. In this review, the detail protocol of real time quantitative PCR technique will be introduced and our recently developed system for exact quantitation of BCR-ABL fusion gene in CML is going to be described.

Related Organizations
Keywords

Base Sequence, Fusion Proteins, bcr-abl, Reproducibility of Results, Polymerase Chain Reaction, Sensitivity and Specificity, Fluorescence, Leukemia, Myelogenous, Chronic, BCR-ABL Positive, Humans, DNA Probes, DNA Primers, Fluorescent Dyes

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Powered by OpenAIRE graph
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selected citations
These citations are derived from selected sources.
This is an alternative to the "Influence" indicator, which also reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Citations provided by BIP!
popularity
This indicator reflects the "current" impact/attention (the "hype") of an article in the research community at large, based on the underlying citation network.
BIP!Popularity provided by BIP!
influence
This indicator reflects the overall/total impact of an article in the research community at large, based on the underlying citation network (diachronically).
BIP!Influence provided by BIP!
impulse
This indicator reflects the initial momentum of an article directly after its publication, based on the underlying citation network.
BIP!Impulse provided by BIP!
19
Average
Top 10%
Top 10%
gold