TY - JOUR AB - The present study aimed to employ an in vivo strategy for the establishment of multi‑copy gene constructs of human DNA topoisomerase I (hTopI) using the pPIC3.5K vector in Pichia. The clones with multi‑copy inserts (His+ transformants) that were able to survive in the highest concentration of Geneticin® were found to express the highest expression level of total protein and exhibited the highest target enzyme activity. The highest level of total protein found was 1.76 mg/ml in GS115‑pPIC3.5K‑hTop1, which was resistant to 1.00 mg/ml Geneticin at 48 h of incubation. The highest enzyme activity of hTopI was also observed in the culture expressed by GS115‑pPIC3.5K‑hTopI, which was resistant to 1.00 mg/ml Geneticin® (19.7x104 Ul/OD600). On the whole, the present study provides information regarding the production of target protein from recombinant Pichia using only a shaker flask system, which can be further developed as an in‑house resource for screening potential anticancer agents. AD - Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia (USM), 11800 Penang, Malaysia AU - Fadzil,Nur,Adila AU - Lim,Shern,Kwok AU - Chew,Ai,Lan AU - Khoo,Boon,Yin DA - 2022/09/01 DO - 10.3892/wasj.2022.167 IS - 5 JO - World Acad Sci J KW - gene copy number protein expression enzyme activity DNA topoisomerase I Pichia pastoris PY - 2022 SN - 2632-2900 2632-2919 SP - 32 ST - Multiple gene copy number increases total protein expression and enzyme activity of DNA topoisomerase I in Pichia pastoris T2 - World Academy of Sciences Journal TI - Multiple gene copy number increases total protein expression and enzyme activity of DNA topoisomerase I in Pichia pastoris UR - https://doi.org/10.3892/wasj.2022.167 VL - 4 ER -