Facile means for quantifying microRNA expression by real-time PCR.
By Rui Shi, Vincent Chiang. BioTechniques, 2005
720 citations recorded by the source corpus.
MicroRNAs (miRNAs) are 20-24 nucleotide RNAs that are predicted to play regulatory roles in animals and plants. Here we report a simple and sensitive real-time PCR method for quantifying the expression of plant miRNAs. Total RNA, including miRNAs, was polyadenylated and reverse-transcribed with a poly(T) adapter into cDNAs for real-time PCR using the miRNA-specific forward primer and the sequence complementary to the poly(T) adapter as the reverse primer. Several Arabidopsis miRNA sequences were tested using SYBR Green reagent, demonstrating that this method, using as little as 100 pg total RNA, could readily discriminate the expression of miRNAs having asfew as one nucleotide sequence difference. This method also revealed miRNA tissue-specific expression patterns that cannot be resolved by Northern blot analysis and may therefore be widely useful for characterizing miRNA expression in plants as well as in animals.
Subfields: Cancer Research, Molecular Biology
View Paper: https://www.semanticscholar.org/paper/e1d0706295ef59e96adcc8d81bc213f6013cd284
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Citations in the corpus
- Genetic unmasking of an epigenetically silenced microRNA in human cancer cells. (2007, methodology): “Real-time PCR analysis of miRNAs was done as previously described (15).”
- Extensive post-transcriptional regulation of microRNAs and its implications for cancer. (2006, methodology): “Mature miRNA PCR was performed according to the high-stringency protocol of Shi and Chiang (2005) except the reverse primer Mir-qPCR-3-3 (5 -GCGAGCA CAGAATTAATACGACTCAC-3 ) was used in conjunction with an exact sequence-specific primer to each miRNA.”
- Serum MicroRNAs Are Promising Novel Biomarkers (2008, methodology): “RNA was subjected to a polyadenylation reaction as described previously [ 17 ].”
- MicroRNAs accurately identify cancer tissue origin (2008, methodology): “One microgram of total RNA was subjected to polyadenylation reaction as described befor”