pCOLADuet-1 is designed for the coexpression of two target genes from a single plasmid. The vector encodes two multiple cloning sites (MCS) each of which is preceded by a T7 promoter, lac operator, and ribosome binding site (rbs). MCS-1 encodes the six-amino acid His•Tag sequence for the creation of a N-terminal fusion and MCS2 encodes the 15 amino acid S•Tag™ peptide after the last restriction site for the creation of a C-terminal fusion if desired. Genes inserted into MCS-1 can be sequenced using the ACYCDuetUP1 Primer and DuetDOWN1 Primer. Genes inserted into MCS-2 can be sequenced using the DuetUP2 Primer and T7 Terminator Primer. The vector has the COLA replicon from ColA(1) and the kanamycin resistance gene. This vector can be transformed into the same cell with plasmids containing compatible origins of replication and drug resistance genes for coexpression of up to 8 target genes.