The article presents a program called RNAhybrid, which is designed to predict multiple potential binding sites of microRNAs (miRNAs) in large target RNAs. The program finds the energetically most favorable hybridization sites of a small RNA in a large RNA, avoiding intramolecular base pairings. For large targets, the algorithm has linear time complexity, allowing for efficient searching of many long targets. The statistical significance of predicted targets is assessed using extreme value statistics of normalized minimum free energies, Poisson approximation for multiple binding sites, and effective numbers of orthologous targets in comparative studies. The method was applied to predict Drosophila miRNA targets in 3'UTRs and coding sequences, identifying known and new targets with significant statistical support. The program RNAhybrid, along with its accompanying programs RNAcalibrate and RNAeffective, is available for download and as a web tool on the Bielefeld Bioinformatics Server.The article presents a program called RNAhybrid, which is designed to predict multiple potential binding sites of microRNAs (miRNAs) in large target RNAs. The program finds the energetically most favorable hybridization sites of a small RNA in a large RNA, avoiding intramolecular base pairings. For large targets, the algorithm has linear time complexity, allowing for efficient searching of many long targets. The statistical significance of predicted targets is assessed using extreme value statistics of normalized minimum free energies, Poisson approximation for multiple binding sites, and effective numbers of orthologous targets in comparative studies. The method was applied to predict Drosophila miRNA targets in 3'UTRs and coding sequences, identifying known and new targets with significant statistical support. The program RNAhybrid, along with its accompanying programs RNAcalibrate and RNAeffective, is available for download and as a web tool on the Bielefeld Bioinformatics Server.