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  • pTWIN-MBP1 Vector

    Discontinued Date

    11/01/2012
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    Categories:
    Discontinued Products

    Description

    pTWIN-MBP1 has the E. coli maltose binding protein (MBP) (2) fused between the modified Ssp DnaB (3) and Mxe GyrA inteins (4). The presence of the chitin binding domain from Bacillus circulans (5,6) facilitates purification. The pTWIN-MBP1 vector permits the isolation of a circular MBP species (1). The double-stranded vector is 8,518 base pairs in length.

    pTWIN-MBP1 contains two mini-inteins, one derived from the Synechocystis sp DnaB intein (154 amino acids) (7) and the other from the Mycobacterium xenopi GyrA intein (198 amino acids) (8).

    Advantages and Features

    Features

    • A pBR322 derivative.
    • Expression of the fusion gene is under the control of the T7 promotor (9) and is regulated by IPTG due to the presence of a lacI gene.
    • Expression requires an E. coli host that carries the T7 RNA Polymerase gene [e.g., T7 Express Competent E. coli (High Efficiency) (NEB #C2566), BL21(DE3) Competent E. coli (NEB #C2527) and derivatives].
    • Origin of DNA replication from the bacteriophage M13 allows for the production of single-stranded DNA by helper phage superinfection of cells bearing the plasmid.
    • Ampicillin resistance.

    Properties and Usage

    Affinity Tag

    Other

    Storage Temperature

    -20°C

    References

    1. Evans, T.C., Benner, J. and Xu, M.-Q. (1999). The cyclization and polymerization of bacterially expressed proteins using modified self-splicing inteins. J. Biol. Chem. 274, 18359-18363.
    2. Guan, C., Li, P., Riggs, P.D. and Inouye, H. (1988). Vectors that facilitate the expression and purification of foreign peptides in Escherichia coli by fusion to maltose-binding protein. Gene. 67, 21-30.
    3. Mathys, S., Evans, T.C., Chute, I.C., Wu, H., Chong, S., Benner, J., Liu, X.-Q. and Xu, M.-Q. (1999). Vectors that facilitate the expression and purification of foreign peptides in Escherichia coli by fusion to maltose-binding protein. Gene. 231, 1-13.
    4. Evans, T.C., Benner, J. and Xu, M.-Q. (1998). Semisynthesis of cytotoxic proteins using a modified protein splicing element. Protein Sci. 7, 2256-2264.
    5. Chong, S., Mersha, F.B., Comb, D.G., Scott, M.E., Landry, D., Vence, L.M., Perler, F.B., Benner, J., Kucera, R.B., Hirvonen, C.A., Pelletier, J.J., Paulus, H. and Xu, M.-Q. (1997). Single-column purification of free recombinant proteins using a self-cleavable affinity tag derived from a protein splicing element. Gene. 192, 271-281.
    6. Watanabe, T., Ito, Y., Yamada, T., Hashimoto, M., Sekine, S. and Tanaka, H. (1994). The role of the C-terminal domain and type III domains of chitinase A1 from Bacillus circulans WL-12 in chitin degradation. J. Bacteriol. 176, 4465-4472.
    7. Wu, H., Xu, M.-Q. and Liu, X.-Q. (1998). Protein trans-splicing and functional mini-inteins of a cyanobacterial DnaB intein. Biochem. Biophys. Acta. 1387, 422-432.
    8. Telenti, A., Southworth, M., Alcaide, F., Daugelat, S., Jacobs, W.R. Jr. and Perler, F.B. (1977). The Mycobacterium xenopi GyrA protein splicing element: Characterization of a minimal intein. J. Bacteriol. 179, 6378-6382.
    9. Dubendorff, J.W. and Studier, F.W. (1991). Controlling basal expression in an inducible T7 expression system by blocking the target T7 promoter with lac repressor. J. Mol. Biol. 219, 45-59.

    Supporting Documents

    Material Safety Datasheets

    The following is a list of Material Safety Data Sheets (MSDS) that apply to this product to help you use it safely. The following file naming structure is used to name these document files: [Product Name] MSDS. For international versions please contact us at info@neb.com.

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    Application Notes