More and more antibiotic analogs are biosynthesized by combinatorial biological techniques. Eleven FK506 analogs are biosynthesized through replacement of allylmalonyl group and 4,5-dihydroxycyclohexa-1,5-dienecarboxylic acid group with heterologous building blocks, and replacement of allylmalonyl biosynthetic elements with heterologous modular biosynthetic elements. These studies will expand our understanding of heterologous building blocks and modular biosynthetic elements and may help us generate more new antibiotic analogs.