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Selection of reference genes in Thermothelomyces fergusii: insights from genomic and expression analysis in response to different carbon sources

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Argitaratua izan da:BMC Genomics vol. 26 (2025), p. 1-15
Egile nagusia: Waheed, Abdul
Beste egile batzuk: Chen, Yi, Hafiz Muhammad Rizwan, Muhammad Adnan, Ma, Xuekun, Liu, Gang
Argitaratua:
Springer Nature B.V.
Gaiak:
Fungi
Genes
Carbon sources
Biological properties
Ranking
Performance evaluation
Molecular biology
Carbon
Correlation coefficients
Correlation coefficient
Biological samples
Cellobiose
Cellulase
Gene expression
Industrial applications
Cellulose
Glucose
Genomic analysis
Algorithms
Thermophilic fungi
Reverse transcription
Environmental
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Laburpena:BackgroundRNA expression analysis using reverse transcription qPCR requires reference genes for accurate data normalization. However, some reference genes can show significant variability in specific biological samples, raising concerns about the reliability of this method. This study assessed the expression levels of 12 reference genes in Thermothelomyces fergusii during spore and mycelium cultivation under different carbon source media (avicel, cellobiose, and glucose) using RT-qPCR. T. fergusii is a thermophilic fungus known for its abundance of cellulase and industrial applications. Quantifying fungal gene expression using the RT-qPCR method is essential for investigating the molecular biology of T. fergusii.ResultsThe stability of 12 reference genes in T. fergusii was evaluated using RefFinder, geNorm, NormFinder, BestKeeper, and delta Ct techniques. The robustness of the chosen reference gene was verified using the LPMO AA9-4 gene. Analysis of the different methods showed variations in the results of reference genes under different sugar treatments. Consistently, EF1, GAPDH, and 18 S-3 were identified as reliable reference genes by the geometric mean of ranking values from Pearson correlation coefficient (r) analysis of pooled data, ranking values of RefFinder in pooled data, and NormFinder of pooled data. However, BT-3, ACT-3, and 18 S-2 exhibited inconsistency as reference genes.ConclusionThese results emphasize the importance of verifying reference genes in specific experimental models and provide valuable recommendations for selecting reliable reference genes in RT-qPCR. These findings have broad applicability for gene expression investigations involving T. fergusii.
ISSN:1471-2164
DOI:10.1186/s12864-025-11772-x
Baliabidea:Health & Medical Collection