PSF-CMV-NH2-FLAG®-EKT-NCOI - N-TERMINAL FLAG® TAG PLASMID OGS96-5UG

NACRES:

NA.85

This plasmid adds a FLAG epitope tag to the N-terminus of a protein that is encoded within the multiple cloning site. This tag allows the detection and purification of a tagged protein using antibodies raised against the FLAG epitope. The FLAG tag coding sequence is DYKDDDDK. There is an enterokinase cleavage site (DDDDK) within the FLAG tag sequence that can be used to remove it from a purified protein if required. 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.
Cloning in a gene: This vector has been designed to allow the addition of a peptide tag to the end of a protein of interest using standard cloning techniques.Multiple Cloning Site Notes:
There is a start codon in the NcoI site can be removed by digestion with KpnI if required. The MCS for gene insertions extends from NotI to XbaI however the tag resides between the NotI and HindIII sites. There are Shine-Dalgarno sequences and KOZAK sequences aligned with the start codon of the peptide tag.
The ClaI to NheI sites have other functions such as adding C-terminal peptide tags second promoters or IRES expression components. The BsgI and BseRI restriction sites cleave within the stop codon in the XbaI site and allow the retrospective fusion of C-terminal peptide tags sequences if the stop codon is placed in this position.
Sequence
Analysis Note
FLAG is a registered trademark of Sigma-Aldrich Co. LLC

form	buffered aqueous solution
mol wt	size 4247 bp
conjugate	FLAG® tagged
Origin of replication	pUC (500 copies)
Peptide cleavage	no cleavage
Peptide tag location	N-terminal
Promoter	Promoter name: CMV Promoter activity: constitutive Promoter type: mammalian
bacteria selection	kanamycin
reporter gene	none
shipped in	ambient
storage temp.	20°C

RIDADR	NONH for all modes of transport
Flash Point F	Not applicable
Flash Point C	Not applicable