Normalization of reverse transcription quantitative-PCR with housekeeping genes in rice

Normalization of reverse transcription quantitative-PCR with housekeeping genes in rice Reverse transcription followed by real-time quantitative polymerase chain reaction (RT Q-PCR) is useful for the systematic measurement of plant physiological changes in gene expression. The validity of using 18S rRNA and three housekeeping genes, glyceraldehyde-3-phosphate dehydrogenase, actin, and tubulin, was tested as a reference of RT Q-PCR. Under various growth stages of etiolated seedlings, different cultivars, and various times after UV-irradiation treatment, expression level of 18S rRNA correlated with total RNA suggesting the uniformity of RT Q-PCR efficiencies among samples. Relative expressions of housekeeping genes varied among samples and independently of experimental conditions, up to two-fold, signifying generally constant fraction of mRNA in total RNA. Results indicate 18S rRNA was the most reliable reference gene for RT Q-PCR of total RNA. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Biotechnology Letters Springer Journals

Normalization of reverse transcription quantitative-PCR with housekeeping genes in rice

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Publisher
Springer Journals
Copyright
Copyright © 2003 by Kluwer Academic Publishers
Subject
Chemistry; Biotechnology; Applied Microbiology; Bioorganic Chemistry; Biochemistry, general; Microbiology
ISSN
0141-5492
eISSN
1573-6776
D.O.I.
10.1023/A:1026298032009
Publisher site
See Article on Publisher Site

Abstract

Reverse transcription followed by real-time quantitative polymerase chain reaction (RT Q-PCR) is useful for the systematic measurement of plant physiological changes in gene expression. The validity of using 18S rRNA and three housekeeping genes, glyceraldehyde-3-phosphate dehydrogenase, actin, and tubulin, was tested as a reference of RT Q-PCR. Under various growth stages of etiolated seedlings, different cultivars, and various times after UV-irradiation treatment, expression level of 18S rRNA correlated with total RNA suggesting the uniformity of RT Q-PCR efficiencies among samples. Relative expressions of housekeeping genes varied among samples and independently of experimental conditions, up to two-fold, signifying generally constant fraction of mRNA in total RNA. Results indicate 18S rRNA was the most reliable reference gene for RT Q-PCR of total RNA.

Journal

Biotechnology LettersSpringer Journals

Published: Jun 9, 2004

References

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