miRNA Profiling on High-Throughput OpenArray™ System

Abstract

Micro RNA (miRNAs) are a class of 17–25 nucleotides noncoding RNAs that have been shown to have critical functions in a wide variety of biological processes. Measuring quantity of miRNAs in tissues of different physiological and pathological conditions is an important first step to investigate the functions of miRNAs. To this date, the number of identified miRNA consists of around 850 different species, and more sequence-predicted miRNA genes are awaiting experimental confirmation. The need for high-throughput technologies allowing to profile all known miRNAs with power similar to microarray and precision/specificity of qPCR is evident. The example of such system based on high-density array of nanoliter PCR assays is described here. Functionally equivalent to a microtiter plate, a single OpenArray™ nanoplate makes possible to do up to 3,072 real-time PCRs at a single experiment. Methods for miRNA profiling using the dual-label probe chemistry (Taqman®) are outlined in this chapter, and experimental data illustrating system performance are provided.