Abstract
Background Pichia pastoris is a methylotrophic species of yeast, which means that it can grow on methanol as its sole carbon and energy source [1]. The Pichia expression system has several advantages: short length of the oligosaccharide chains added to proteins, correct folding and very high cell densities [2]. P. pastoris vectors are designed for homologous integration into either AOX I locus, one of the two homologous AOX I genes present in this species or his4 locus. Gene insertion events at the AOX I (GS115) loci arise from a single crossover events between the loci and any of the three AOX I regions in the vector, the AOX I promoter, the AOX I transcription termination region (TT), or sequences even further downstream of AOX I (3′ AOX I) [3]. In either GS115 gene insertion events at his4 locus arise from a single crossover event between his4 locus in the chromosome and His4 gene of the vector. This results in the insertion of one or more copies of the vector in his4 locus [3].
Highlights
Pichia pastoris is a methylotrophic species of yeast, which means that it can grow on methanol as its sole carbon and energy source [1]
Recombinants clones P pastoris GS115 with pPG Vector integrad in the AOX locus were transformad with pPAmy vector, both previouly contructed in our laboratory
The supernatants were collected by centrifugation and subjected to enzyme activity assay by Fuwa and DNS methods
Summary
Pichia pastoris is a methylotrophic species of yeast, which means that it can grow on methanol as its sole carbon and energy source [1]. Methods Recombinants clones P pastoris GS115 (phenotype His +/Muts) with pPG Vector (pPIC9/ glucoamylase cDNA of Aspegillus awamori) integrad in the AOX locus were transformad with pPAmy vector (pPIC9 vector/a-amylase gene of Bacillus licheniformis), both previouly contructed in our laboratory. The pPAmy was linearizad by digesting with SacI enzyme to HIS4 locus integration. Transformation was carried out by electroporation of freshly prepared competent cells.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
