Protocol for high-throughput miRNA profiling of the rat brain

Abstract

MicroRNA (miRNA) are a class of small, noncoding RNA molecules that participate in the posttranscriptional regulation of gene expression. These molecules modulate gene expression by either repressing translation or inducing mRNA degradation, and collectively may regulate as much as two-thirds of the transcriptome. By affecting gene regulation, miRNAs are likely to be involved in most biological processes, with small variations in miRNA expression able to modify the expression of numerous genes within a biological network. It is likely therefore that miRNA dysregulation underlies many of the molecular changes observed in a range of neuropathology. Certainly, the identification of brain-specific miRNA is indicative of their important role in both the function and development of the central nervous system (CNS). The brain is the most complex tissue in the mammalian organism and animal models such as the rat provide an affordable and reliable means to investigate the role of miRNA in relation to structure, function, and behavior. Here we present a systematic approach to examining genome-wide expression of miRNA in the rat brain. This method involves the isolation and quantification of total RNA, the generation of biotin- labelled RNA fragments, as well as whole genome expression microarrays. High-throughput miRNA profi ling is a powerful experimental tool that enables insight into the genetic underpinnings of physiological and pathological processes. It has vast implications in identifying specific miRNA involved in developmental and pathological processes including synaptic plasticity and mental illness.