Golden Gate (GG) cloning assembles DNA fragments in a defined order to create a novel expression unit. The six entry modules contain a promoter, N-terminal tag, gene of interest, C-terminal tag, terminator, and a resistance marker. Expression units are assembled into one of three vectors that serve as both destination vectors for GG and entry vectors for Gateway (GW). The three GW entry vectors (pEN-L4-AG-R1, pEN-L1-AG-L2, and pEN-R2-AG-L3) are flanked by attR and attL sites for GW recombination. GW recombination with a destination vector results in an expression clone with three, ordered expression cassettes. The ccdB and chloramphenicol resistance (CmR) genes between the cloning sites provide positive and negative selection, respectively.