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Mammalian

OG101 - pSF-CMV-BetaGal

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Full Plasmid Details (PDF)

Sequence (txt)

This plasmid is compatible with all of the other DNA sequences available on this site, which are all sold in the same backbone. This flexibility allows you to create any vector you require, with a variety of functions, simply by compiling the relevant sections from our product range.

Vector: pSF-CMV-BetaGal

Product Code: OG101

Quantity Provided: 5µg

Size (bp): 7430

Bacterial Antibiotic Selection: Ampicillin

Origin and Compatibility: pUC high copy, derived from pBR322   

Copy Number: 500-700 copies per cell

Promoter: Cytomegalovirus (CMV) immediate early promoter

Purpose: A versatile expression vector for the expression on the beta galactosidase enzyme from the CMV immediate early promoter.

Transcription Termination: 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.

C-Terminal Fusions: This plasmid contains the beta galactosidase gene in the main multiple cloning site. Any of our C-terminal coding tags and sequences can be fused to the C-terminus of the beta galactosidase gene in a single cloning step. This can be performed by cutting the vector with BsgI, or BseRI, and any other downstream restriction site (such as NheI). Cleaving any of our C-terminal tag plasmids (with the suffix 1 at the end of the name) with the same restriction sites will create compatible ends. When ligated together the stop codon of the beta galactosidase gene will be removed, and the C-terminal tag will be in frame with the beta galactosidase coding sequence.

Intellectual Property Status: According to our IP-friendly policy, this plasmid is sold free of reach-through rights on any derivatives you may create.

Quality Validation: This vector has been demonstrated to express beta galactosidase to high levels under the CMV promoter. The start codon of the reporter gene was positioned within the NcoI site and the stop codon was positioned within the XbaI site. Expression was validated in the A549 lung carcinoma cell line.  

Quality Sequencing Analysis: This vector has been fully sequenced using primers F1-F10. The sequences and details of these primers are available through our website.

Genetic Modifications to standard parts:

  • CMV promoter – an NcoI site has been ablated in the CMV promoter. This change does not reduce expression.
  • AmpR cassette – This AmpR resistance region is not a wild type sequence, it has been modified to remove all restriction sites that conflict with the SnapFast system and to retain a high level of antibiotic resistance. It has been validated in E.coli.
  • Beta Galactoidase – The beta gal gene has been modified to remove ClaI, EcoRV, SacI and PciI restriction sites.   

Restriction site notes:

  • Bgl2, SwaI and PmeI each cut the vector at two sites positioned to flank the promoter, origin, and KanR, respectively.
  • BsgI and BseRI recognise non-palindromic DNA sequences and cleave upstream of their recognition sites, within the stop codon that is found inside the XbaI restriction site. The beta galactosidase stop codon is also cleaved by these two sites, allowing fusions to be made.