PSF-CMV-RAT-KAPPA LC - RAT KAPPA LIGHT CHAIN ANTIBODY PLASMID OGS525-5UG

This vector is for the production of kappa antibody light chains for rat (Rattus rattus). It has been engineered so that when the vector is cut with the restriction enzyme BseRI it produces an overhang from the first codon of the constant region. This allows antibody light chain variable regions to be seamlessly ligated into this vector to create full length rat kappa light chain antibody expression cassettes. As part of this vector range we also provide plasmids for the production of both human and mouse antibodies as well as vectors for the creation of Heavy IgG chain and Lambda light chain rat antibodies (named pSF-CMV-Rat-IgG HC and pSF-CMV-Rat-Lambda respectively)
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.
The plasmid encodes a constant region an antibody in the main multiple cloning site positioned so that it can be cleaved to produce an overhang that allows seamless fusion with a variable region from any antibody. This allows you to create full length antibody genes with no cloning scars.
To enable this immediately upstream of the constant region coding sequence there is a BseRI restriction site. This is a type-IIS restriction enzyme that binds in one position (CAGCAG) and then cleaves a specific number of nucleotides away from the binding site regardless of the sequence at the cleavage point. We use this site in all of our antibody expression cassettes in the same position. In this plasmid cutting with BseRI will result in an overhang consisting of the first two nucleotides of the first codon of the constant region. This means that any variable region with the same overhang at its 3 prime end can be ligated into this plasmid when used in conjunction with any 5 prime site (NotI-NcoI). To add this overhang the variable region must be PCR amplified to contain any of the following sites at its 3 prime end: BseRI BsgI BtsI or BsrDI. By using this system it allows antibody variable regions to PCR amplified and fused to any of our constant region plasmids without having to re-synthesise the entire antibody expression cassette each time.
Sequence
Analysis Note

form	buffered aqueous solution
mol wt	size 4576 bp
Origin of replication	pUC (500 copies)
Peptide cleavage	no cleavage
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