Heterologous Biosynthesis of Cordycepin in Aspergillus oryzae
[Objective]This study aims to synthesize cordycepin by constructing engineered Aspergillus oryzae strains that heterologously express key genes for cordycepin synthesis.[Method]The uidine/uracil and histidine diauxotrophic strain of Aspergillus oryzae AoΔpyrGΔHisB served as the background,and overexpressed the key gene CmCns1-CmCns3 for cordycepin synthesis using the Agrobacterium-mediated transformation technique.The subcellular localization of CmCns1 and CmCns2 within A.oryzae was visualized through fluorescence microscopy.High-performance liquid chromatography(HPLC)was used to determine the content of cordycepin in transgenic A.oryzae.Additionally,glycine and adenine,both precursors in the synthesis of cordycepin,were introduced into the fermentation broth to investigate their impact on cordycepin synthesis in A.oryzae.[Result]The CmCns1 and CmCns2 from Cordyceps militaris were localized in lipid droplets in A.oryzae.Cordycepin synthesis achieved through the overexpression of CmCns1 alone,the co-expression of CmCns1 and CmCns2,and the co-expression of CmCns1-CmCns3 in A.oryzae,and the highest extracellular production of cordycepin reached 37.74 μg/mL after 48 h of fermentation.The addition of glycine and adenine to the fermentation broth of A.oryzae could not effectively increase the content of cordycepin.[Conclusion]This study successfully achieved heterologous expression of synthetic cordycepin genes in A.oryzae.
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