tac Vectors for High Level Bacterial Expression
Bacterial tac promoter-based vectors allow expression, detection and purification of recombinant FLAG and MAT (Metal Affinity Tag) fusions in E. coli. These vectors offer a choice of periplasmic (+ompA) or cytoplasmic expression of amino- or carboxy-terminal tagged fusions. tac expression vectors confer ampicillin resistance for easy selection of positive transformants. Additionally, the vectors contain the T1T2 transcriptional terminator, pMB1(derivative of pBR322) origin of replication, the f1 origin, and the lacI gene for repression of the tac promoter. Vectors utilizing the strong tac promoter (a hybrid of the E. coli trp and lac promoters) offer protein expression levels in excess of 10 mg/L of culture when using IPTG as a de-repressor. These vectors can be used to express protein in any established E. coli expression host.
The MAT tag or Metal Affinity Tag (HNHRHKH) has been created for purification of recombinant MAT fusion proteins using HIS-Select Nickel and Cobalt Affinity Gels. HIS-Select products allow for highly selective purification of histidine-tagged fusion proteins such as MAT fusions. Many of our newest vectors make use of the MAT tag, often in combination with the well-known FLAG tag. MAT tag containing vectors are offered in formats for N-terminal or C-terminal tagging.
FLAG-SHIFT vectors are available for sequences where the amino-terminus has not been precisely defined. These vectors contain a "shift" sequence that allows expression of an insert in all three reading frames, thus without regard to the reading frame in which it was originally cloned.
The recognition sequence for enterokinase, Asp-Asp-Asp-Asp-Lys, is found at the C-terminal end of the FLAG epitope tag. Removal of FLAG is possible in all fusion proteins containing an N-terminal FLAG sequence. Dual tag fusion proteins may also be cleaved with enterokinase for removal of one or more tags, depending on the position of FLAG in the protein sequence.
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