Identification and functional validation of target genes from a large and complex wheat genome are challenging. However, recent progress in genomic sequencing and assembly can facilitate rapid cloning of wheat genes. We applied these advancements and have developed a low-coverage, long-read PacBio contig assembly of wild emmer wheat (Triticum turgidum ssp. dicoccoides). Together with bulked segregant analysis sequencing (BSA-Seq) and analyses of EMS-derived susceptible mutants this assembly has led to the identification of the novel stripe rust resistance gene Yr84. The Yr84 originates from wild emmer wheat accession PI 487260, which is highly resistant to several tested stripe rust isolates. Through BSA-Seq analysis the gene was localized to chromosome arm 1BS, while further fine-mapping allowed narrowing of resistance to a 975 kb physical region. To facilitate characterization of the functional allele, we generated a low-coverage (~10X) PacBio assembly of PI 487260. Whole-genome, short-read resequencing of EMS-derived susceptible mutant lines of PI 487260 revealed seven lines that carry mutations in a single candidate gene with an NLR (nucleotide-binding domain leucine-rich repeat-containing) structure. A whole-genome blast search for Yr84 sequence in wild emmer Zavitan, durum wheat Svevo and bread wheats from the 10+ Genome project revealed Yr84 copies on chromosomes 1B and 2B in all these assemblies, though non-functional due to SNPs and structural variations. We have developed a functional molecular marker and screened a tetraploid wheat diversity panel (~500 genotypes) for the presence of Yr84. This analysis revealed that the functional allele is rare among wild emmer wheat accessions and was completely absent in the domesticated emmer and durum wheat landrace gene pools screened in this study. The combined approach of low-coverage long-read genome assembly and BSA-Seq was an effective strategy to rapidly characterize Yr84 and to develop functional markers that can support breeding.