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| |  | Cat. No. 71341 |
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| Coexpression of multiple target proteins in E. coli | | | The Duet vectors are T7 promoter expression vectors, each designed to coexpress two target proteins in E. coli. Use the Duet vectors together or with existing pET, pCDF, pRSF, and pETcoco™ recombinants with compatible replicons and antibiotic resistance to coexpress up to eight proteins in the same cell (Novy 2002, Held 2003).
Compatible replicons and drug resistance genes
The Duet vectors are designed with compatible replicons and drug resistance genes for effective propagation and maintenance of four plasmids in a single cell:
• pETDuet™ carries the ColE1 replicon and bla gene (ampicillin resistance)
• pACYCDuet™ carries the P15A replicon and cat gene (chloramphenicol resistance)
• pCDFDuet™ carries the CloDF13 replicon and aadA gene (streptomycin/ spectinomycin resistance)
• pRSFDuet™-1 carries the RSF1030 replicon and kan gene (kanamycin resistance).
• pCOLADuet™ carries the COLA replicon from ColA and the kan gene (kanamycin resistance).
Each vector carries two expression units, each controlled by a T7lac promoter for high-level protein expression. Each promoter is followed by a ribosome binding site and multiple cloning region. A T7 terminator follows the second multiple cloning sequence. The multiple cloning regions have restriction sites that facilitate the cloning of two genes and the transfer of inserts from other pET constructs. The Duet vectors are designed with the option of producing native unfused proteins or fusions to His•Tag® and S•Tag™ sequences for detection and purification of protein complexes. | | | Novy, R., et al. 2002. inNovations 15, 2. Held, D., et al. 2003. inNovations 18, 4. | | | | | |
pRSFDuet™-1 is designed for the coexpression of two target proteins. The vector encodes two multiple cloning sites (MCS) each of which is preceded by a T7 promoter, lac operator, and ribosome binding site (rbs). The vector also carries the pRSF1030 replicon (also known as NTP1), lacI gene, and kanamycin resistance marker. This vector can be transformed into the same cell with pETDuet™-1, pACYCDuet™-1, and/or pCDFDuet™-1 for the coexpression of up to eight target proteins. ORFs inserted into MCS-1 can be sequenced using the ACYCDuetUP1 Primer and DuetDOWN1 Primer. ORFs inserted into MCS-2 can be sequenced using the DuetUP2 Primer and T7 Terminator Primer. |
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