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Overlapping Promoter Library Designed for Rational Heterogenous Expression in Cordyceps militaris

Summary

This research developed a new method to control gene expression in the medicinal mushroom Cordyceps militaris by stacking multiple copies of genetic switches called promoters. This advancement helps scientists better engineer beneficial compounds in mushrooms. Impacts on everyday life: – Improved production of medicinal compounds from mushrooms for healthcare – More efficient and sustainable manufacturing of natural therapeutic products – Advancement of techniques to enhance beneficial properties of edible mushrooms – Potential development of new pharmaceutical products from fungi

Background

Cordyceps militaris is an edible and medicinal fungus widely accepted in East Asia that has potential as a cell factory for producing adenosine analogs. While gene editing techniques and genome modeling have advanced, the diversity of DNA elements in C. militaris remains too limited to achieve rational heterogeneous expression for metabolic engineering studies.

Objective

To develop and evaluate an overlapping promoter library strategy in C. militaris using PtrpC promoters to achieve quantitative expression of multiple target genes. The study aimed to expand promoter engineering capabilities in edible mushrooms through BioBricks assembly of overlapping promoters.

Results

Both mRNA transcription and GFP expression levels gradually increased with the copy number of overlapping promoters, peaking at 7 copies. Compared to single promoter strains, the transcription level increased 63-fold in strains with 7 promoter copies. No significant differences were found in biomass or morphological characteristics between engineered and wild-type strains.

Conclusion

This study successfully demonstrated the first application of overlapping promoter strategy in C. militaris, providing proof-of-concept for fungi synthetic biology and establishing a general method for promoter engineering in edible mushrooms. The approach enables rational balancing of multiple genes without overusing promoter sources.
  • Published in:Microbial Cell Factories,
  • Study Type:Laboratory Research,
  • Source: 10.1186/s12934-022-01826-0
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