Wheat is one of the world's most important crops, providing 20% of global calorific intake. A constant threat to production of this cereal crop are rust diseases caused by fungal pathogens of the Puccinia genus. Rust diseases are most economically and sustainably managed by breeding resistant wheat crops containing rust resistance genes. However constant evolution of new, aggressive rust pathogen requires new sources of resistance to be identified in unimproved landraces and wild relatives of wheat. Transfer of resistance from wild species into domesticated wheat by conventional breeding is time consuming, expensive and often impeded by linkage drag of undesirable traits linked to resistance genes of interest. Isolation of rust resistance genes enables their direct transfer into elite wheat cultivars by transformation or alternatively these cloned gene sequences can be used as perfect markers to facilitate conventional breeding. By exploiting current advances in DNA sequencing coupled with gene capture technology we have developed a rapid resistance gene cloning methodology called Mutagenesis, Resistance gene enrichment and Sequencing (MutRenSeq) which was used for isolation of two globally important stem rust resistance genes. This new approach greatly accelerates the isolation of these agronomically important resistance genes from the large and complex hexaploid wheat genome and will enable new biotechnological disease resistance approaches to be undertaken; such as stacking multiple resistance genes at a single locus. This technology is currently being tested in other important crop species.