Prokaryotic expression is mature and easy to be operated, but when it comes to the proteins of eukaryotic organism, error folding is occurred frequently since glycosylation and folding partner are involved. Fortunately,the expression system ofpichia pastorisis complementary to this situation to a certain extent. Relying on powerful synthetic biology technology platform and industry-leading codon optimization softwareNGTMCodon, Synbio Tech. provides you high-quality and considerate yeast expression protein services with more than 10 years’ experience in protein expression and purification, which can effectively save your cost and time.
1.Expression vector construction (optional)
First, according to your specific requirements, we will analyze the gene sequence of protein of interest. Through the industry-leading NGTMCodon optimization software, the amount of gene expression can be increased by several to tens of times. Then, gene synthesis was conducted and the recombinant plasmids are verified by sequencing to see whether the sequences are 100% correct. Synbio Tech. provides the service of directly cloning genes into commercial expression vectors such as methanol-induced pPICZα、pPIC9K or component pGAPZα, etc.. In addition, you can also choose the peculiar yeast special expression vector of Synbio Tech.to carry on the expression test.
Delivery contents: 1 tube of freeze dried plasmid (2~5 µg/tube) and 1 tube of glycerol containing the plasmid TOP10
Delivery TAT: 10-20 business days
2.Linearized vector andelectricity transforming into pichiapastoris
You can also give us a constructed recombinant plasmid (please confirm the codon of the sequence is optimized). For those verified recombinant plasmids, we choose suitable enzymes to carry out the linearization and adopt the electric transfer methodto transform them into the pichiapastoris. After the screening, we select at least 6 colonies for expression laboratory scale test.
Delivery contents: screening report of positive colonies
Delivery TAT: 5 business day
3.Optimization of expression conditions
During the process oflaboratory scale test，we will optimize and testthe culture conditions which are suitable for various strains, including temperature, culture medium, inducer type, induction time, induction quantity and other factors.It is tacitly approved thatwe use SDS-PAGE or Western blotto (optional, the customer provides a varified first antibody) to analyze the expression effect.
Delivery contents: A copy of laboratory scale test verification report
Delivery TAT: 5 business days
Amplified cultivation,expression and purification interest protein
To optimize and obtain the strains with appropriate expression conditions and use 1L or greater culture scale to induce recombinant yeast expression. After the induction, one step or multi-step purification is conducted.we use SDS-PAGE to verify the purification andWestern blotto (optional, the customer provides a varified first antibody) to verify the accuracy.
Delivery contents: 1-10 mg recombinant proteins and purification report data
Delivery TAT: 10-15 working days
For proteinswith tags, youcan choose remove service (optional).
Protein post processing (optional)
For those proteins with further special requirements, Synbio Tech. provides you a variety of post processing options, including protein tag removal after purification, filtration sterilization, endotoxin removal, freeze-dry, N-terminal sequencing and other services.Our experienced engineers can provide tailor-made post servicesfor your special needs.