Product Name: pSF-CMV-NH2-His-EKT-NcoI
Product Code: OG211
Size (bp): 4256 bp
Bacterial Antibiotic Selection: KanR
Origin and Compatibility: pUC high copy derived from pBR322
Bacterial Copy Number: 500-700 per cell
Promoter: Cytomegalovirus (CMV) immediate early promoter
This vector adds a His epitope tag to the N-terminus of a protein that is encoded within the multiple cloning site. This tag allows the purification of a tagged protein by binding to metal matrices such as nickel or cobalt. There is an enterokinase cleavage site (DDDDK) immediately downstream of the His tag that can be used to remove the His tag from a purified protein. It cleaves after the lysine residue.Promoter Expression Level:
This plasmid contains the mammalian CMV promoter to drive gene expression. We have tested all of our mammalian promoters in a range of cell types and CMV is consistently the strongest in those we have studied. However there are many reports of the CMV promoter demonstrating silencing by methylation in long-term culture. For this reason we stock a range of other promoters that are compatible with this plasmid and are available on request.
This plasmid contains three alternative transcription terminators for mammalian bacterial and bacteriophage (T7) expression. This means that only the promoter needs to be changed to alter the expression system you are using. We sell multiple promoters that can be used in each of these systems. The presence of each terminator does not reduce expression in the alternative systems.
The his tag is in frame with the ATG start codon that is within the NcoI restriction site allowing fusion with any genes that we sell in the MCS that contain an NcoI at the 5 prime. This tag is positioned immediately upstream of the NcoI site to minimise the amount of extra amino acids that are added to the N-terminus of your gene.
The start codon that is within the NcoI restriction site is in frame with and adjacent to the coding sequence for the His/EKT peptide tag. Shine-Dalgarno sequences and KOZAK sequences are positioned correctly at the start of the His signal peptide. The ClaI to NheI sites have other functions such as adding peptide tags or IRES elements. The BsgI and BseRI restriction sites cleave within the stop codon in the XbaI site and allow fusion with our C-terminal peptide tags provided that your genes stop codon is also positioned here.