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Quantitative analysis of gene expression by reverse transcription polymerase chain reaction and capillary electrophoresis with laser-induced fluorescence detection

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Quantitative analysis of gene expression by reverse transcription polymerase chain reaction and capillary electrophoresis with laser-induced fluorescence detection

Content Provider	Springer Nature Link
Author	Richards, Mark P. Poch, Stephen M.
Copyright Year	2002
Abstract	There has been a dramatic expansion of DNA sequence information compiled over the past several years for a variety of eukaryotic and prokaryotic genomes. Accompanying this increase in knowledge of genomic structure and organization has been a growing interest in studying the function of individual genes including regulation of their expression. A number of methods such as Northern blotting, ribonuclease protection assay, and hybridization arrays have been developed to analyze gene expression at the transcriptional (mRNA) level. Although quantitative estimates of mRNA transcripts can be obtained from each of these methods, oftentimes they lack sufficient sensitivity or the methodology is too costly or too labor-intensive to be applied to the analysis of a large number of samples. The most sensitive method for analyzing gene expression at the mRNA level involves the combination of reverse transcription and polymerase chain reaction (RT-PCR). However, in order to provide accurate quantitative estimates of gene expression, a rapid and efficient method is required for separation and detection of the double-stranded DNA (dsDNA) products of RT-PCR. Recent advances in capillary electrophoresis with laser-induced fluorescence detection (CE/LIF) have made this method suitable for the automated analysis of large numbers of RT-PCR samples. An overview of the application of CE/LIF to quantitative analysis of gene expression by RT-PCR is presented along with selected protocols and examples. Both relative-quantitative (RQ) and quantitative-competitive (QC) approaches to RT-PCR are discussed in conjunction with the use of CE/LIF for rapid and accurate quantitative analysis of PCR products.
Starting Page	19
Ending Page	37
Page Count	19
File Format	PDF
ISSN	10736085
Journal	Molecular Biotechnology
Volume Number	21
Issue Number	1
e-ISSN	15590305
Language	English
Publisher	Humana Press
Publisher Date	2002-01-01
Publisher Place	Totowa
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Neurosciences Neurology
Content Type	Text
Resource Type	Article
Subject	Molecular Biology Biochemistry Bioengineering Applied Microbiology and Biotechnology Biotechnology

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