Product Name: pSF-AOX1-Zeo
Product Code: OG3584
Size (bp): 5646 bp
Bacterial Antibiotic Selection: KanR
Origin and Compatibility: pUC high copy derived from pBR322
Bacterial Copy Number: 500-700 per cell
Promoter: Yeast (strong) constitutive triosephosphate isomerase (TPI1) gene
This vector is designed for the expression of a gene of interest (GOI) in the methylotrophic yeast, Pichia pastoris. Once the GOI has been cloned into the MCS, the plasmid is then linearised and transformed into an appropriate P. pastoris strain by electroporation. The expression construct becomes integrated into the genome. Expression of the GOI is induced by addition of methanol to the culture medium.The vector contains a Zeocin resistance marker for selection of integrants in any Pichia strain. Increasing the Zeocin concentration also enables selection of multicopy integration.Promoter Expression Level:
This promoter is inducible by methanol and exhibits high expression under these conditions. In the absence of methanol the promoter is tightly regulated and shows only basal levels of expression.
This plasmid contains the poly-adenylation signal from the Alcohol Oxidase 1 Promoter (AOX1) gene from Pichia pastoris to terminate transcription.
This plasmid has been designed to be compatible with a range of cloning techniques. The multiple cloning site contains a range of standard commonly used restriction sites for cloning. Using these sites genes can be inserted using standard cloning methods with DNA ligase. Other methods such as ligase independent cloning (LIC) Gibson Assembly InFusionHD or Seamless GeneArt can also be used and because all of our plasmids are based on the same backbone the same method can be used for cloning into all of our catalogue vectors.Multiple cloning site notes:
There are a few important sites within the MCS. These include the NcoI site the XbaI site and the BsgI and BseRI sites. The NcoI site contains a start codon that is immediately downstream of both a Kozak and Shine-Dalgarno ribosomal binding site. These allow for optimal positioning of genes when the start codon is placed in this location. If this is not required and you wish to use a downstream site for gene cloning you can remove the NcoI site by cleaving the plasmid with KpnI.
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.
This product is part of our SnapFast™ plasmid range, for more information on the intellectual property status of this plasmid and the terms of our licences please click here.