Abstract: Cultural and nutritional conditions were optimized for Indole Acetic Acid (IAA) production by Rhizobium spp. isolated from root nodules of Crotalaria juncea, C. retusa, C. laburnifolia, C. verrucosa and C. alata. The isolates produced maximum amount of IAA after 72 h of incubation and at 2.5 mg mL^-1 L-tryptophan concentration. The effect of different carbon and nitrogen sources on IAA production were also studied and it revealed that, mannitol and L-glutamic acid were the best promoters for IAA production. Addition of cell wall affecting agents increased the IAA production over controls. Among the five isolates of Crotalaria species, maximum amount of IAA was produced by isolate from C. retusa. The compound from Rhizobium sp. from C. retusa was extracted, purified and structurally confirmed as IAA.