pPIC3.5K Vector

pPIC3.5K Vector

pPIC3.5K is a vector used to identify the integration of multiple gene copies into the Pichia pastoris genome. The pPIC3.5K vector is 9004 bp in size. In the multiple cloning site region, it contains five unique restriction enzyme sites: BamHI, SnaBI, EcoRI, AvrII, and NotI. It is used for intracellular expression. During vector construction, it is necessary to include an ATG start codon and a Kozak sequence (Cavener and Stuart, 1991; Kozak, 1987; Kozak, 1990). The HIS4 gene is used for selection in Pichia pastoris. To insert a gene into the AOX1 locus of GS115 and KM71, a SacI-linearized plasmid is used (GS115 generates a His⁺ Mut⁺ genotype, KM71 generates a His⁺ MutS genotype). To insert a gene into the HIS4 locus of GS115 and KM71, a SalI-linearized plasmid is used (GS115 generates a His⁺ Mut⁺ genotype, KM71 generates a His⁺ MutS genotype). A gene can also be inserted into the AOX1 gene of GS115 using a BglII-linearized plasmid, producing yeast with a His⁺ MutS genotype. The inserted gene must include the ATG start codon.

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