Many disease resistance genes have been introgressed into wheat from its wild relatives. However, reduced recombination within the introgressed segments hinders cloning of the introgressed genes. Here, we report the identification of wheat powdery mildew resistance gene Pm13 originated from from Aegilops longissima using a method combining physical mapping with radiation-induced chromosomal aberrations and transcriptome sequencing analysis of ethyl methanesulfonate (EMS)-induced loss-of-function mutants. Pm13 encodes a kinase fusion protein, designated MLKL-K, with an N-terminal domain of mixed lineage kinase domain-like protein (MLKL_NTD domain) and a C-terminal serine/threonine kinase domain bridged by a brace. The resistance function of Pm13 is validated by transient and stable transgenic complementation assays. Transient over-expression analyses in Nicotiana benthamiana leaves and wheat protoplasts reveal that the fragment Brace-Kinase_122-476 of MLKL-K is capable of inducing cell death, which is dependent on a functional kinase domain and the three α-helices in the brace region close to the N-terminus of the kinase domain.