Sexual reproduction in flowering plants involves double fertilization, the union of two sperm from pollen with two sex cells in the female embryo sac. Modern plant breeders increasingly seek to circumvent this process to produce doubled haploid individuals, which derive from the chromosome-doubled cells of the haploid gametophyte. Doubled haploid production fixes recombinant haploid genomes in inbred lines, shaving years off the breeding process1. Costly, genotype-dependent tissue culture methods are used in many crops2, while seed-basedin vivodoubled haploid systems are rare in nature3and difficult to manage in breeding programmes4. The multi-billion-dollar maize hybrid seed business, however, is supported by industrial doubled haploid pipelines using intraspecific crosses toin vivohaploid inducer males derived from Stock 6, first reported in 1959 (ref.5), followed by colchicine treatment. Despite decades of use, the mode of action remains controversial6,7,8,9,10. Here we establish, through fine mapping, genome sequencing, genetic complementation, and gene editing, that haploid induction in maize (Zea mays) is triggered by a frame-shift mutation inMATRILINEAL(MTL), a pollen-specific phospholipase, and that novel edits inMTLlead to a 6.7% haploid induction rate (the percentage of haploid progeny versus total progeny). Wild-type MTL protein localizes exclusively to sperm cytoplasm, and pollen RNA-sequence profiling identifies a suite of pollen-specific genes overexpressed during haploid induction, some of which may mediate the formation of haploid seed11,12,13,14,15. These findings highlight the importance of male gamete cytoplasmic components to reproductive success and male genome transmittance. Given the conservation ofMTLin the cereals, this discovery may enable development ofin vivohaploid induction systems to accelerate breeding in crop plants.