|pAB-6xHis™ plasmid transfer vector is designed for expression of proteins with N-terminal polyhistidine tag. A PCR fragment encoding a protein of interest can be cloned in lieu of the polyhedrin promoter and polyhistidine tag using any of the unique restriction endonuclease sites (SmaI, XbaI, EcoRI, NotI, PstI, BglII) located in the MCS. Proteins are expressed under control of strong very late polyhedrin promoter in the polyhedrin site of baculovirus DNA. Affinity purification of the expressed proteins can be achieved using Ni-NTA (nickel-nitrilotriacetic acid) resin (QUIAGEN). If desired, polyhistidine tag can be cleaved with thrombin at thrombin cleavage site (LVPRGS) encoded at nucleotides 4174-4191 of the vector. Unique SpeI site can be used to obtain 6xHis-tagged proteins without the thrombin site. Unique BamHI site can be use to obtain proteins without polyhistidine tag. Positions of phF and mR primers, which are useful for sequencing the insert/vector junctions, are shown on the DNA sequence as arrows.|
The above plasmid transfer vector is compatible with linearized baculovirus DNA from several other suppliers, e.g. BacMagic™, BacMagic™-2, BacMagic™-3, BacVector™-1000, BacVector™-2000, BacVector™-3000 (Novagen); BaculoGold™, BaculoGold™ bright (BD Pharmingen); flashBACGOLD™, flashBACULTRA™, flashBAC™ (OET); BacPAK6™ (Clontech); Sapphire™ (Orbigen). They are not compatible with Bac-to-Bac® and BaculoDirect™ DNA from Invitrogen. BacMagic™, BacMagic™-2, BacMagic™-3, BacVector™-1000, BacVector™-2000, BacVector™-3000 are trademarks of Novagen; BaculoGold™, BaculoGold™ bright are trademarks of BD Pharmingen; flashBACGOLD™, flashBACULTRA™, flashBAC™ are trademarks of OET; BacPAK6™ is a trademark of Clontech; Sapphire™ is a trademark of Orbigen, Bac-to-Bac® and BaculoDirect™ are trademarks of Invitrogen.
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