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PinPoint Xa Protein Purification System

by Promega
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  • Overview
  • Protocols
  • Specifications
  • Resources
  • Express and Purify In Vivo-Biotinylated Fusion Proteins

    Express and purify fusion proteins that are biotinylated in vivo. The DNA coding for the protein of interest is cloned into a PinPoint™ Vector downstream of a sequence encoding a peptide that becomes biotinylated in vivo. Biotinylated fusion proteins are produced in E. coli and are affinity-purified using the SoftLink™ Soft Release Avidin Resin. This proprietary resin allows elution of the fusion protein under nondenaturing conditions. The PinPoint™ Vectors feature the encoded endoproteinase Factor Xa (pronounced "ten a") proteolytic site that provides a way to separate the purification tag from the native protein, and the vectors carry a convenient multiple cloning region for ease in construction of fusion proteins.

    The system contains vectors in all possible sense reading frames, an avidin-conjugated resin, Streptavidin-Alkaline Phosphatase, a purification column and biotin. The PinPoint™ Xa Control Vector contains the chloramphenicol acetyltransferase (CAT) gene provided as a means of monitoring protein expression, purification and processing conditions. The system generally yields 1–5mg of protein per liter of culture.

    Additional features include the flexibility to use SoftLink™ Resin for release of the fusion protein under nondenaturing conditions and presence of the tac promoter, which allows tightly regulated expression.

    Applications

    • Purification of milligram amounts of expressed protein.
    • Production of novel bioreagents.

    Reference

    1. Cress, D. et al. (1993) Promega Notes 42, 2–7.
    0923VA-W

    PinPoint™ Xa-1 Vector map.

    Sequence Information

    PinPoint™ Xa-1 Vector GenBank® Accession Number U47626.

    PinPoint™ Xa-2 Vector GenBank® Accession Number U47627

    PinPoint™ Xa-3 Vector GenBank® Accession Number U47628.

  • Protocols

    Complete Protocol

    Download PDF

    PinPoint™ Xa Protein Purification System Technical Manual

    PDF (480 KB)

  • Certificate of Analysis

    Lookup Certificate of Analysis

    Storage Conditions

    +2C TO +10C

    Use Restrictions

    For Research Use Only. Not for Use in Diagnostic Procedures.

  • Resources

    Citations