Comprehensive evaluation and validation of optimal reference genes for normalization of qPCR data in different...

Area of Science:

• Molecular Biology
• Genomics
• Animal Science

Background:

• Quantitative real-time PCR (qPCR) is crucial for validating gene expression data.
• Accurate qPCR necessitates normalization using stable reference genes.

Purpose of the Study:

• To identify optimal reference genes for qPCR in various goat tissues.
• To ensure reliable gene expression profiling in caprine studies.

Main Methods:

• Evaluated 18 candidate reference genes across 10 caprine tissues.
• Utilized RefFinder, integrating NormFinder, GeNorm, BestKeeper, and ΔCt algorithms.
• Assessed gene expression stability using computational analysis.

Main Results:

• ACTB, PPIB, and B2M identified as the most stable reference genes.
• RPL19, RPS15, and RPS9 determined to be the least stable.
• Validated selected genes for accurate gene expression profiling.

Conclusions:

• The geometric average of ACTB, PPIB, and B2M serves as an effective normalization factor.
• These reference genes are recommended for gene expression studies in goat tissues.
• Establishes a foundation for robust molecular research in goats.