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Puromycin Selection Plasmids

OG136 - pSF-CMV-Ub-Puro-SV40 Ori Sbfl

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Product Code: OG136

Size (bp): 6342

Bacterial Antibiotic Selection: Kanamycin

Origin and Compatibility: pUC high copy, derived from pBR322   

Copy Number: 500-700 copies per cell

Promoter: Cytomegalovirus (CMV) immediate early promoter and the Human Ubiquitin (Ub) promoter

Purpose: 
This vector contains a puromycin (puro) resistance expression cassette under the Ub promoter. It also contains the Simian Virus 40 (SV40 origin of replication. This can be used to increase transgene signal following transient transfection. It is also reported that it is possible to make cell lines containing the plasmid as a multi-copy episome, however, we have not validated it for this purpose. The Ub puro cassette is flanked by AscI sites in this plasmid and is not designed to be split up. If you would like the Puromycin gene alone, or the Ub promoter alone, please see the mammalian selection gene section, or the promoter section, on our website. This vector can also be used to force recombination into the genome of mammalian cells not expressing SV40 large T antigen following transfection and puromycin selection, although this method can be inefficient. If cells are expressing the SV40 large T antigen, those that integrate the plasmid would likely experience chromosomal instability because the large T antigen would initiate DNA replication from within the host chromosome.

Inserting a Gene: Each restriction site in this vector has a purpose and allows the insertion of specific pre-designed DNA sequences. To insert a gene, we recommend using any sites between (and including) the HindIII and XbaI sites. By doing this you will be able to insert most of our other products upstream, or downstream, of your gene. Click here to read more about the purpose of each restriction site.

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 Sequencing Analysis: This vector has been fully sequenced using primers F1-F10. These are available through our website using the sequencing primers link on the left hand lower menu.

Restriction site notes:

  • Bgl2, KpnI, SwaI, AscI and PmeI each cut the vector at two sites positioned to flank the promoter, start codon, origin, Ubiquitin Puromycin cassette, and the 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.

Genetic Modifications to standard parts:

  • CMV promoter – an NcoI site has been ablated in the CMV promoter. This change does not reduce expression.
  • KanR cassette – This KanR 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.
  • Puromycin resistance gene - The puro gene has been modified to remove a StuI site and an EagI site to ensure compatibility with all of our products. These are all conservative changes and do not change the amino acid sequence.   
  • Ub promoter - The Ubiquitin (Ub) promoter in this vector has been modified to remove BseRI, StuI and SpeI sites. These changes do not reduce expression.
  • SV40 Origin - The SV40 origin has been modified to remove NcoI, BamHI, ClaI, StuI and BseRI sites. These changes to not alter the core SV40 origin region and do not alter its function.