Selection of reference genes for RT-qPCR analysis in developing chicken embryonic ovary

Area of Science:

• Reproductive biology
• Developmental biology
• Molecular genetics

Background:

• Accurate gene expression normalization requires stable internal reference genes.
• Identifying reliable reference genes is crucial for studying chicken embryo ovary development and oocyte meiosis initiation.

Purpose of the Study:

• To evaluate and confirm the stability of candidate reference genes for gene expression analysis in chicken embryonic ovaries.
• To identify the most suitable internal reference genes for studying molecular regulation of oocyte meiosis initiation.

Main Methods:

• Evaluated expression profiles of six candidate genes (Actb, Cvh, Dazl, Eef1a, Gapdh, Rpl15) in chicken embryonic ovaries (E12.5-E20.5).
• Assessed gene stability using GeNorm, NormFinder, BestKeeper, and Comparative ΔCt methods.
• Combined results using RefFinder to rank reference gene stability.

Main Results:

• Eef1a and Rpl15 were identified as the most stable reference genes.
• Cvh was found to be the least stable reference gene.
• Marker genes (Stra8, Scp3, Dmc1) for oocyte meiosis entry were successfully normalized using Eef1a and Rpl15.

Conclusions:

• Eef1a and Rpl15 are the most suitable internal reference genes for chicken embryo ovary development studies.
• These validated reference genes will facilitate further research into the molecular mechanisms of oocyte meiosis initiation.