Screening and identification of miRNAs related to epidermal color in Jianbai Xiang pigs based on transcriptome sequencing

【Objective】The purpose of the study was to sequence the transcriptome of black and white epidermal tissues of Jianbai Xiang pig， to screen out miRNA that regulated melanin production， and to provide a theoretical basis for further study on the regulatory mechanism of miRNA on epidermal color formation of Jianbai Xiang pig. 【Method】The black and white epidermal tissues of Jianbai Xiang pigs were used as experimental materials for transcriptome sequencing. DESeq2 was used to screen differentially expressed miRNAs with|log₂ Fold Change|≥1 and false discovery rate（FDR） ≤ 0.05 as the standard. The target genes of differentially expressed miRNAs were predicted by miRanda and RNAhybrid，and the enrichment analysis of KEGG signaling pathway was performed by GO functional annotation. The interaction network of differentially expressed miRNAs， target genes and KEGG signaling pathway was constructed， and the results of transcriptome sequencing were verified by real-time fluorescence quantitative PCR. 【Result】A total of 280 known miRNAs and 618 new miRNAs were identified from the black and white epidermis of Jianbai Xiang pig， and 17 differentially expressed miRNAs were screened. Ten miRNAs were up-regulated and seven miRNAs were down-regulated in the white epidermis compared with the black epidermis. A total of 1327 target genes were predicted and obtained. GO functional annotation showed that in biological processes， the differentially expressed miRNA target genes were mainly involved in epithelial development， cell proliferation and catabolic processes. In cell components and molecular functions， they were mainly involved in cytoplasm， cytoplasmic vesicles， ATP binding and catalytic activity. The enrichment results of KEGG signaling pathway showed that the target genes were enriched in phosphatidylinositol signaling system， melanin production and cytochrome P450 signaling pathways. Among them， ssc-miR-615 target genes included CALM3， CREBBP， FZD5 and DVL2， and their enrichment pathways were all related to epidermal pigmentation. The enrichment pathways of ssc-miR-221-3p target gene TYRP1 and ssc-miR-224 target gene RAB23 were related to melanin production. Seven differentially expressed miRNAs and target genes TYRP1 and RAB23 were selected for real-time fluorescence quantitative PCR verification. The results showed that the expression changes of differentially expressed miRNAs were consistent with the changing trend of transcriptome sequencing analysis. The expression of TYRP1 and RAB23 in the black epidermis of Jianbai Xiang pigs was extremely significantly higher than that in the white epidermis （P<0.01）. 【Conclusion】 The ssc-miR-615， ssc-miR-221-3p and ssc-miR-224 are candidate genes affecting the epidermal color of Jianbai Xiang pig， which may have an important effect on the formation of melanin in the epidermis of Jianbai Xiang pig.