MicroRNAs (miRNAs) are similar to 22 nt non-coding RNA molecules that usually function as endogenous repressors of target genes. Many biological processes depend on faithful miRNA expression and miRNA profiling has revealed dysregulation of many miRNAs in neurological, and cardiovascular diseases, and in cancer. Despite this finding, most studies have focused on the function of single miRNAs or miRNA clusters. To better address physiologically relevant collaborative miRNA interactions, we developed a simple and flexible platform which expresses several miRNAs that have different genomic locations from a single transcript using endogenous pre-miRNA sequences. As a proof of principle we cloned the miR-34 tumor suppressor family and showed that the miR-34a/34b/34c vector expresses each miRNA at similar levels to individual miRNA containing vectors. Moreover, the miR-34a/34b/34c vector suppressed cell growth more than the individual miRNA vectors. We expect that this platform will be invaluable as a tool to study the complex and synergistic interactions of aberrantly expressed miRNAs in human diseases and may have applications for use in gene therapy. (C) 2011 Elsevier Inc. All rights reserved.