Hereditary stability of Thermomyces lanuginosus lipase gene in recombinant Pichia pastoris strain

Abstract:

[Background] hereditary stability of recombinant strain is the prerequisite for high-level and stable expressing, which determines the success of industrial applications. [Objective] hereditary stability of Pichia pastoris recombinant strain containing heterologous Thermomyces lanuginosus lipase gene was studied. [Methods] recombinant strain was subcultured for 15 generations and its stability judged from the following aspects: colony morphology, cell morphology, hydrolytic activity of lipase, DNA sequence and integrated copy number of the target gene. [Results] colony and cell morphology of the 1st, 5th, 10th and 15th generations of strains remained the same, as well as the molecular weight of the target protein and the target gene sequence. It was also found that after pass-generation culture for 5 times, the integrated copy number of lipase gene was greatly reduced and then stabilized at about 7 in the 10th and 15th generations, while the relative hydrolytic activity of lipase reached up to 90%. [Conclusion] high-level expression of the lipase is achieved by moderate gene copy number. this recombinant strain has a good hereditary stability and is feasible for large-scale production.

Key words:Hereditary stability, Pichia pastoris, Lipase, Recombinant strain, Industrial application