Abstract：Tree frog (Rhacophorus) antimicrobial peptide Cathelicidin is a kind of small molecular peptide with antibacterial activity, which has potential application value in medical treatment, animal husbandry and some other aspects, but its eukaryotic expression is rarely reported. In this study, tree frog antimicrobial peptide Cathelicidin gene was synthesized based on the preferential codon usage of Pichia pastoris, then ligated to the expression vector pPIC9K and transformed into P. pastoris GS115 by electroporation. The positive transformants containing multi-copy gene insertions were screened using high concentration G418 and confirmed by PCR and reverse transcription PCR (RT-PCR). The recombinant Cathelicidin was induced for 72 h with 1.0% methanol, and the supernatant of culture medium was collected for in vitro antibacterial activity detection and toxicity analysis. The results showed that Cathelicidin was expressed successfully in P. pastoris with antibacterial activity against gram-positive bacteria (Staphylococcus aureus) and gram-negative bacteria (Escherchia coli), the minimal inhibitory concentration were (1.175±0.002) and (2.35±0.001) μg/mL respectively, and it was no hemolysis under the minimum inhibitory concentration. This study provides basic data support for industrial production of a new tree frog antimicrobial peptide.
王莲哲, 刘士俊, 刘佳乐, 洪军. 树蛙抗菌肽Cathelicidin在毕赤酵母中的表达及抑菌活性分析[J]. 农业生物技术学报, 2021, 29(1): 67-72.
WANG Lian-Zhe, LIU Shi-Jun, LIU Jia-Le, HONG Jun. Expression of Tree Frog (Rhacophorus) Cathelicidin Peptide in Pichia pastoris and Its Antibacterial Activity Analysis. 农业生物技术学报, 2021, 29(1): 67-72.
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