Inhibiting miRNA in Caenorhabditis elegans using a potent and selective antisense reagent

We have developed a new class of fluorescently labelled antisense reagents to inhibit miRNAs in developing worms. These reagents were synthesized by conjugating dextran with 2'-O-methyl oligoribonucleotide. The dextran-conjugated antisense reagents can be conveniently introduced into the germline of adult hermaphrodites and are transmitted to their progeny, where they efficiently and specifically inhibit a targeted miRNA in different tissues, including the hypodermis, the vulva and the nervous system. We show that these reagents can be used combinatorially to inhibit more than one miRNA in the same animal.This class of antisense reagents represents a new addition to the toolkit for studying miRNA in C. elegans. Combined with numerous mutants or reporter stains available, these reagents should provide a convenient approach to examine genetic interactions that involve miRNA, and may facilitate studying functions of miRNAs, especially ones whose deletion strains are difficult to generate.See related research article: http://jbiol.com/content/9/3/20 webciteMicroRNAs (miRNAs) are single strand RNA molecules ~ 21-23 nucleotides long that play important roles in many biological processes through regulating gene expression [1]. In animal cells, miRNAs act primarily by inhibiting mRNA translation and/or stability through a process involving partial complementary base-pairing with sequences at the 3'-untranslated region (3' UTR). Numerous miRNAs have been identified. To study their functions, antisense reagents against miRNAs have been developed as a reverse genetics tool. Synthetic oligonucleotide analogues, including 2'-O-methyl oligoribonucleotides [2], locked nucleic acids [3], 2'-O-methoxyethyl oligoribonucleotides [4], and morpholinos [5], have been tested. These antisense nucleotide analogues have been used to knock down miRNAs in cultured cells [2-4] and in live animals including zebrafish [5], D. melanogaster [6] and mice [7].Caenorhabditis elegans has long been used