Product Name: pSF-CAG-BetaGal-Ub-Puro
Product Code: OG506
Size (bp): 10450 bp
Bacterial Antibiotic Selection: AmpR
Origin and Compatibility: pUC high copy derived from pBR322
Bacterial Copy Number: 500-700 per cell
Promoter: CAG synthetic mammalian promoter and Ubiquitin promoter / Chicken Beta Actin (CBA) promoter / Human Ubiquitin Promoter
This plasmid is designed for creating beta galactosidase stable cell lines or the transient expression on beta galactosidase. It contains the strong constitutive mammalian CAG promoter to drive the expression of the beta galactosidase reporter and a puromycin resistance gene to allow for stable cell line selection.Promoter Expression Level:
This plasmid contains the mammalian CAG promoter which is a synthetic composite of the CMV immediate early enhancer followed by the CBA promoter and the rabbit beta globin intron. The Chicken beta actin contains a CpG island that can help to keep the promoter active for longer in stable culture when compared to the CMV promoter. The ubiquitin promoter that is driving expression of the puromycin gene demonstrates a high level of expression in most cell types. It demonstrates similiar levels of expression to the commonly used EF1-Alpha promoter.
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.
This plasmid contains a gene within the main multiple cloning site (NotI-ClaI). Any plasmid that we sell where the gene is this configuration will be located in the exact same position in relation to the start and stop codon of the gene. The only exceptions to this rule are fusions proteins where the fusion gene may be positioned at the front or end of the MCS to allow gene fusion.
By positioning all of our genes in the same location it allows them to be transferred between plasmids using the same cloning method and restriction sites regardless of the plasmid being used from our product range. Inserting a new gene into this plasmid should be easily possible using a range of standard restriction enzyme sites that flank the gene currently in the vector.Multiple cloning site notes:
In the multiple cloning site there are two important restriction sites called BsgI and BseRI sites. These sites both cut the DNA at the same position and cleave the stop codon of the gene in the multiple cloning site in this plasmid thereby producing a TA overhang. This overhang is compatible with any of our peptide or reporter fusion tag plasmids also cut with either of these enzymes. This allows seamless C-terminal fusions to be made with the gene in this multiple cloning site using a single cloning step from our C-terminal peptide and reporter tag product range. Normally the easiest method is to clone the C-terminal tag from our other plasmid products into this plasmid using BsgI or BseRI and the downstream ClaI restriction site.
BseRI and BsgI sites are non-palindromic and cleave a defined number of bases away from their binding sites. This allows them to cut the upstream stop codon in the gene in this plasmid regardless of the gene sequence.