Identification and Functional Validation of ACSL1 and FABP3 as Muscle-Related Genes Screened by Transcriptomics in Crossbred Duroc × Berkshire × Diannan Small-Eared Pigs

<b>Background:</b> Crossbreeding strategies that combine the growth performance of Western pig breeds with the meat quality traits of Chinese indigenous breeds have garnered considerable interest. Duroc pigs are known for their high growth efficiency but have relatively low intramuscular fat (IMF) content. In contrast, native breeds like the Diannan Small-Eared pig exhibit superior pork quality with higher IMF levels. This study aimed to compare the muscle growth characteristics and molecular mechanisms between Duroc × Landrace × Yorkshire (DLY) and Duroc × Berkshire × Diannan Small-Eared (DBD) pigs. <b>Methods:</b> The longissimus dorsi tissue of 210-day-old DLY and DBD pigs was collected for analysis. HE staining assessed muscle fiber characteristics, IMF content was measured, and ELISA quantified muscle-derived growth and development-related factors. Transcriptome sequencing was conducted, followed by differential gene expression analysis, Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and protein-protein interaction (PPI) analyses. Functional validation of key genes was performed in C2C12 cells. <b>Results:</b> DBD pigs exhibited significantly larger muscle fiber diameter and higher IMF content compared to DLY pigs. IGF1 and GH levels were elevated in DBD pigs. Transcriptome analysis identified 185 upregulated and 102 downregulated genes, with enrichment in pathways including PI3K-Akt, MAPK, FoxO, and cGMP-PKG signaling. <i>ACSL1</i> and <i>FABP3</i> were functionally validated, showing promotion of differentiation and inhibition of proliferation in C2C12 cells. <b>Conclusions:</b> DBD pigs exhibit superior muscle growth traits and higher IMF content compared to DLY pigs. <i>ACSL1</i> and <i>FABP3</i> may serve as key regulators of muscle development in pigs.