Background: This study explores the potential of discarded male layer embryos as a sustainable and non-GMO cell source for cultivated chicken meat production. The research aims to identify efficient methods for isolating muscle progenitor cells (MPCs) with high proliferative potential by conducting transcriptome analysis on thigh muscle tissues from both male and female chick embryos. Methods: Transcriptome analysis was performed on the thigh muscle tissues of male and female chick embryos, aged 12-13 days, (n = 4 each), to investigate the gene expression profiles and identify strategies for efficiently isolating MPCs. This approach aims to pinpoint techniques that would allow for the selection of MPCs with optimal growth and proliferation capabilities. Results: Using heatmap, hierarchical clustering, and multidimensional scaling (MDS), we found no significant sex-based differences in gene expression, except for the overexpression of the female-specific gene LIPBLL. The expression of muscle stem cell factors, including PAX3, PAX7, and other myogenic regulatory genes, showed no significant variation. However, to recover MPC-rich cells isolated from male thigh muscle, we found that by the pre-plating 7 stage, myogenesis-related genes, MYHs and MUSTN1 were minimally expressed, while the cell cycle arrest gene CDKN1A sharply increased. Conclusions: Our findings suggest that simple cell isolation directly from tissue is a more scalable and efficient approach for cultivated meat production, compared to labor-intensive pre-plating methods, making it a viable solution for sustainable research and resource recycling.

ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
    Reagents and ethics statements
    Sample acquisition and processing
    Isolation of musclec progenitor cells (MPCs)
    Transcriptome analysis
    Statistical analysis
RESULTS
    Transcriptome analysis of male and female thighmuscles and stage-specific cells
    Expression of transcripts related to myogenicregulation factors (MRFs)
    Transcripts expression related to cell lifespan andproliferation
    Expression of MYHs and MUSTN1 involved in skeletalmuscle maturation
DISCUSSION
CONCLUSION
REFERENCES

• Sun A Ock(Animal Biotechnology Division, National Institute of Animal Science, Rural Development Administration, Wanju 55365, Korea) Corresponding author
• Yeongji Kim(Animal Biotechnology Division, National Institute of Animal Science, Rural Development Administration, Wanju 55365, Korea)
• Young-Im Kim(Animal Biotechnology Division, National Institute of Animal Science, Rural Development Administration, Wanju 55365, Korea)
• Poongyeon Lee(Animal Biotechnology Division, National Institute of Animal Science, Rural Development Administration, Wanju 55365, Korea)
• Bo Ram Lee(Animal Biotechnology Division, National Institute of Animal Science, Rural Development Administration, Wanju 55365, Korea)
• Min Gook Lee(Animal Biotechnology Division, National Institute of Animal Science, Rural Development Administration, Wanju 55365, Korea)