The in vivo haploid induction approach offers several advantages compared to the in vitro induction approach and recurrent self-pollination. It is currently the method of choice for inbred line development in many commercial maize breeding programs. Here, we describe the in vivo approach for generation of maternal doubled haploids (DHs). It involves four steps: (1) induction of haploidy by pollinating source germplasm with pollen of a haploid inducer; (2) identification of putative haploid seeds (seeds with a haploid embryo) using a seed coloration marker system; (3) doubling of chromosomes of putative haploids by treating seedlings with a mitotic inhibitor; and (4) verification of putative doubled haploids with a stalk color marker and self-pollination of true doubled haploid plants to multiply their seed.