The continuously increasing genome sequencing data has revealed numerous cryptic pathways, which might encode novel secondary metabolites with interesting biological activities. However, utilization of this hidden potential has been hindered by the observation that many of these gene clusters remain silent (or poorly expressed) under laboratory conditions. Here we present reporter-guided mutant selection (RGMS) as an effective and widely applicable method for targeted activation of silent gene clusters in the native producers. The strategy Lakes advantage of genome-scale random mutagenesis for generation of genetic diversity and a reporter-guided selection system for the identification of the desired target-activated mutants. It was first validated in the re-activation of jadomycin biosynthesis in Streptomyces venezuelae ISP5230, where high efficiency of activation was achieved. The same strategy was then applied to a hitherto unactivable pga gene cluster in Streptomyces sp. PGA64 leading to the identification of two new anthraquinone aminoglycosides, gaudimycin D and E. (C) 2015 International Metabolic Engineering Society. Published by Elsevier Inc. All rights reserved.