Integration Stability of sHBsAg-Multi Expression Cassettes in Pichia pastoris GS115 during Methanol Induction
Hepatitis B is the major health problem worldwide including in Indonesia. Vaccination is the best prevention strategy for the disease. For the purpose of vaccine development and to decrease drug import, production of Hepatitis B Virus (HBV) small surface antigen (sHBsAg) from Indonesian HBV subtype is needed. The recombinant protein production can be conducted by integrating multi expression cassettes of sHBsAg gene in Pichia pastoris chromosome using gene replacement method. Such integration method turns out to allow loss of foreign gene from chromosome by excisional recombination-mediated looping out. This research was aimed to determine integration stability of four copies of sHBsAg expression cassette in P. pastoris GS115 chromosome inducted with 2% methanol in FM22 medium. The methanol induction was conducted twice at 63-h and 75-h. Integration stability determination was conducted qualitatively using PCR and quantitatively using qPCR absolute quantification. A band of 208 bp with similar intensity was observed after amplification of genomic DNA. All samples generated the same Ct value of around 22 with four copies of sHBsAg gene per genome. The result from this experiment shows that integration of four copies of sHBsAg expression cassette in P. pastoris GS115 chromosome is stable during methanol induction.
HAYATI J Biosci article's license is CC-BY-NC. This license lets others distribute, remix, tweak, and build upon author's work, as long as they credit the original creation.
Authors who submit and publish with this journal agree to the following terms:
- Authors retain copyright and grant the journal/publisher non exclusive publishing rights with the work simultaneously licensed under a https://creativecommons.org/
licenses/by-nc/4.0/Attributi on — You must give appropriate credit, provide a link to the license, and indicate if changes were made. You may do so in any reasonable manner, but not in any way that suggests the licensor endorses you or your use.
- Authors can still use their work commercially
- Authors are able to enter into separate, additional contractual arrangements for the non-exclusive distribution of the journal's published version of the work (e.g., post it to an institutional repository or publish it in a book), with an acknowledgement of its initial publication in this journal.
- Authors are permitted and encouraged to post their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work (See The Effect of Open Access).