Agrobacterium rhizogenes -mediated transformation results in the development of hairy roots at the site of infection. The production of hairy roots involves cocultivation of explants with A. rhizogenes and the subsequent selection of hairy roots on hormone-free medium. Hairy roots have many applications for research including secondary product production and for the study of biochemical pathways. In addition, transgenic plants regenerated from hairy roots often show an altered phenotype due to the presence of the rol genes. In this chapter we describe how to produce and grow hairy root cultures, how to regenerate shoots from these hairy roots, and how to conduct molecular analysis of these cultures.