PSF-CMV-PUC18 - CMV PUC18 MCS PLASMID OGS107-5UG

NACRES:

NA.85

A versatile cloning plasmid for the expression of genes in mammalian cells. This plasmid contains the multiple cloning site (MCS) from pUC18 however it has been modified slightly to accommodate some restriction sites in our vector system. These changes are described in the full plasmid details PDF. The use of this MCS instead of the our normal vector MCS will limit the ability to use some of the inserts that we sell that immediately flank or are inserted within the standard MCS. This primarily includes N-terminal tags and signal peptides. This is because these inserts are flanked by restriction sites that are not compatible with the pUC18 MCS. Most other inserts should still be compatible with this plasmid.
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.
Multiple cloning site notes: This vector contains a different multiple cloning site (MCS) from the standard SnapFast vector. The MCS is derived from pUC18. The order of the restriction sites in this vector are almost the same as in pUC18 however the sequence is not. The pUC18 MCS has also been modified as follows: The XbaI site in the pUC18 MCS has been replaced with a SpeI site (these sites produce compatible cohesive ends when cleaved).

• The SbfI and PstI sites have been replaced with an NsiI site (which produces compatible cohesive ends with SbfI and PstI when cleaved).
• The BamHI site that is normally found in all SnapFast vectors (in the downstream fusion MCS) has been replaced with a BclI site because the pUC18 MCS contains this site. BamHI and BclI produce compatible cohesive ends when cleaved although BclI is methylated in this vector and will require growth in a Dam methylase negative bacterial strain (such as JM110 cells) before it can be used.
• 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 the retrospective fusion of coding sequences. These sites are normally only used on genes that we sell in the main multiple cloning site. This concept is explained in more detail on our website using the drop down box on the homepage and selecting: Fuse a coding sequence to the 3 prime end of a gene already in the plasmid.
• 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 the retrospective fusion of coding sequences. These sites are normally only used on genes that we sell in the main multiple cloning site. This concept is explained in more detail on our website using the drop down box on the homepage and selecting: Fuse a coding sequence to the 3 prime end of a gene already in the plasmid.

Sequence
Analysis Note

form	buffered aqueous solution
mol wt	size 4310 bp
Origin of replication	pUC (500 copies)
Peptide cleavage	no cleavage
Promoter	Promoter name: CMV Promoter activity: constitutive Promoter type: mammalian
bacteria selection	ampicillin
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