Reference genes for proximal femoral epiphysiolysis expression studies in broilers cartilage

Area of Science:

• Molecular Biology
• Animal Science
• Genetics

Background:

• Accurate gene expression analysis relies on stable reference genes for normalization.
• Reference gene stability can vary significantly with experimental conditions, necessitating validation.
• Epiphysiolysis is a condition affecting broiler femur development, requiring reliable molecular markers.

Purpose of the Study:

• To identify the most stable reference genes for gene expression studies in broiler femur cartilage.
• To evaluate nine candidate reference genes under normal and epiphysiolysis conditions.
• To provide recommendations for reliable endogenous controls in avian skeletal research.

Main Methods:

• Collected femur articular cartilage from 29 male broilers (35 days old).
• Performed RNA extraction followed by quantitative PCR (qPCR) analysis.
• Utilized GeNorm, BestKeeper, and NormFinder software for gene stability assessment, with RankAggreg for global ranking.

Main Results:

• Evaluated nine candidate reference genes: GAPDH, HMBS, HPRT1, MRPS27, MRPS30, RPL30, RPL4, RPL5, and RPLP1.
• RPLP1 and RPL5 demonstrated the highest stability across the evaluated samples.
• A global ranking confirmed RPLP1 and RPL5 as the most reliable endogenous genes.

Conclusions:

• RPLP1 and RPL5 are recommended as the most reliable reference genes for gene expression studies in broiler femur cartilage.
• These genes are suitable for research involving epiphysiolysis and other femur-related anomalies in broilers.
• Validated reference gene selection enhances the accuracy and reproducibility of gene expression data in avian skeletal research.