Isoflavonoids are a diverse group of secondary metabolites derived from the phenylpropanoid pathway. These compounds are distributed predominantly in leguminous plants and play important roles in plant-environment interactions and human health. Consequently, the biosynthetic pathway of isoflavonoid compounds has been widely elucidated in the past decades. Up to now, most of the structural genes and some of the regulatory genes involved in this pathway have been isolated and well characterized. Nowadays, the protective effects of the legume isoflavonoids against hormone dependent cancers, cardiovascular disease, osteoporosis, and menopausal symptoms have generated considerable interest within the genetic and metabolic engineering fields to enhance the dietary intake of these compounds for disease prevention. Subsequently, there are some great progresses in genetic and metabolic engineering to improve their yields in leguminous and non-leguminous plants and/or microorganisms. Because of the field of flavonoid biosynthesis has been reviewed fairly extensively in the past, this review concentrates on the more recent development in the isoflavonoid branch of phenylpropanoid pathway, including gene isolation and characterization. In addition, we describe the state-of-the-art research with respect to genetic and metabolic engineering of isoflavonoid biosynthesis.