pTYB21 is an E. coli cloning and expression vector (7514 bp) used in the IMPACT™ Protein Purification System which allows the overexpression of a target protein as a fusion to a self-cleavable affinity tag (1,2). It is an N-terminal fusion vector designed for in-frame insertion of a target gene into the polylinker, downstream of the intein tag (the Sce VMA intein/chitin binding domain, 55 kDa)(3,4). This allows the N-terminus of the target protein to be fused to the intein tag. The self-cleavage activity of the intein allows the release of the target protein from the chitin-bound intein tag, resulting in a single column purification of the target protein.
This vector can be used in conjuction with a C-terminal fusion vector to test which fusion construction (N-terminal or C-terminal) maximizes the expression and yield of a target protein. For the fusion of the C-terminus of the target protein to the intein tag, use pTXB1 (NEB #N6707), pTXB3 (NEB #N6708), pTYB1 (NEB #N6701), pTYB2 (NEB #N6702), pTYB3 (NEB #N6703) or pTYB4 (NEB #N6704).
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