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Act1 out of Action: Identifying Reliable Reference Genes in Trichoderma reesei for Gene Expression Analysis

Summary

Scientists studying the fungus Trichoderma reesei, which is used industrially to produce enzymes, found that the reference genes commonly used to measure gene activity (act1 and sar1) are not as reliable as previously thought. By analyzing genetic data from multiple studies, they identified two better reference genes called bzp1 and tpc1 that maintain more consistent expression levels across different growth conditions. Using these better reference genes will help researchers more accurately measure how genes are expressed in this important industrial fungus.

Background

Trichoderma reesei is an important industrial enzyme producer used for cellulase and hemicellulase production. Gene expression analysis via RT-qPCR is fundamental to understanding enzyme regulation, but requires stable reference genes for accurate normalization. The commonly used reference genes act1 and sar1 were selected based on literature reports rather than systematic validation.

Objective

This study aimed to identify and validate alternative reference genes for T. reesei by analyzing publicly available transcriptome datasets and evaluating expression stability under relevant cultivation conditions. The researchers sought to determine whether act1 and sar1 are truly suitable as universal reference genes for RT-qPCR normalization in this organism.

Results

All five newly identified candidate genes (bzp1, tpc1, cue1, ubi1, sas3) demonstrated more stable expression than act1 and sar1. Bzp1 and tpc1 ranked as the most stable reference genes. The study revealed that act1 and sar1 exhibit strain- and condition-dependent expression variability, making them unsuitable as universal reference genes.

Conclusion

The authors recommend using bzp1 and tpc1 in combination for RT-qPCR normalization in T. reesei instead of the traditional act1 and sar1. The study emphasizes the importance of validating reference gene stability for new organisms, strains, and experimental setups to ensure reliable gene expression analysis.
  • Published in:Journal of Fungi (Basel),
  • Study Type:Experimental Validation Study,
  • Source: PMID: 40422730, DOI: 10.3390/jof11050396
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