Pricing & Availability
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|69954-3||Plastic ampoule||40 rxn||
This product has been discontinued.
This kit is available as a convenient supplement to the pET Peptide Expression System 31b, which is designed for expression and purification of target peptides (1). In pET-31b(+), peptide coding sequences are placed downstream of a 125 amino acid (aa) ketosteroid isomerase (KSI) gene and upstream of a His•Tag® sequence. Use of the unique AlwN I cloning site enables unidirectional insertion of coding sequences immediately adjacent to a methionine residue. For high-yield production, small (10–25 aa) peptide coding sequences can be cloned as tandem repeats interspersed with single methionine residues. The KSI-peptide-His•Tag fusion protein is expressed at high levels in the presence of T7 RNA polymerase provided by the recA–, protease deficient BLR(DE3)pLysS host. The fusion protein is purified in one step under denaturing conditions using one of Novagen's His•Bind® metal chelation resins. It is then cleaved with CNBr to release the His•Tag peptide, insoluble KSI, and monomer peptide units each terminating with a homoserine lactone. The purified peptide can be easily converted to an amide or homoserine, or reacted with other compounds, such as analogs of fluorescein and biotin, to produce C-terminal derivatized peptides. The system has been used to make biologically active peptides of 14–67 aa at yields of > 50 mg purified peptide per liter of culture (1).
The AlwN I digested, dephosphorylated vector is ready to accept appropriate inserts containing ...ATG-3′ and 3′-TAC... overhangs on the top and bottom strands, respectively. A Control Insert is provided to verify performance and ensure optimal cloning efficiency.
|References||1. Kuliopulos, A. and Walsh, C.T. (1994) J. Am. Chem. Soc. 116, 4599–4607.|
|Safety Information according to GHS|
|Product Usage Statements|
|Storage and Shipping Information|
|Ship Code||Shipped with Blue Ice or with Dry Ice|
|Avoid freeze/thaw||Avoid freeze/thaw|
|Do not freeze||Ok to freeze|
|1. Kuliopulos, A. and Walsh, C.T. (1994) J. Am. Chem. Soc. 116, 4599–4607.|
|TB053 Academic and Non-profit Laboratory Assurance Letter|
|TB055 pET System Manual|
|TB104 pET Peptide Expression System 31|
|TB098VM pET-31b(+) Vector Map|