Year: 2005
Type: article
Abstract: microRNAs (miRNAs) are small RNAs that regulate translation and hence control a variety of cellular processes in metazoans. The quantitation and identification of miRNAs has been hampered by their small size and low abundance. Here we describe two robust PCR-based assays of miRNA expression based on the original cloning strategy. The non-quantitative PCR method allows detection and identification of miRNAs and we utilise this method in the discovery of a new miRNA (miR-532) in retinoic acid differentiated P19 cells. The second and quantitative method (QM-RT-PCR) is simple and accurate, and uses commonly available technology. Of particular interest is the specificity of this PCR-based technology compared to hybridisation-based methods including arrays and northern blotting. Here we have shown that a single base pair mismatch in the priming sequence results in a two order of magnitude reduction in the amplification of let-7f. These streamlined methods will complement previously described methods and will facilitate analysis of miRNA expression in rare cell populations where the amount of RNA is limited.
Source: PubMed Central
Institutions: The University of Sydney, Centenary Institute
Language: English
Subfield: Cancer Research
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W1543624430