Identification of long non-coding RNAs involved in fat 1 deposition in pigs using two high-throughput sequencing 2 methods 3

: 23 Background: Adipose is an important body tissue in pigs, and fatty traits are critical in pig 24 production. The function of long non-coding RNA (lncRNA) in fat deposition and metabolism has 25 been proven in previous studies. In this study, we focused on lncRNAs associated with fattening 26 traits in pigs. The adipose tissue of six Landrace pigs with either extremely-high or -low backfat 27 thickness ( n high = 3, n low = 3) was collected, after which we performed strand-specific RNA 28 sequencing using biological replicates and pooling methods. 29 Results: A total of 19,631 genes and 2,013 lncRNAs were identified using the coding potential 30 calculator, coding-non-coding index, and Pfam database, including 334 known transcripts and 31 1,679 novel transcripts. Using edgeR, we determined that 220 lncRNAs and 1,512 genes were 32 differentially expressed (|Fold Change| > 2 and false discovery rate < 0.05) between the two 33 groups in biological replicate RNA sequencing (RNA-seq), and 127 lncRNAs and 2,240 genes 34 were differently expressed in pooling RNA-seq. Further Kyoto Encyclopedia of Genes and 35 Genomes and Gene Ontology enrichment analysis of the differentially expressed genes found that 36 some of the genes were involved in several key pathways related to fat development. After 37 targeting gene prediction, we determined that some cis-target genes of the differentially expressed 38 lncRNAs play an important role in fat deposition. For example, ACSL3 is cis-targeted by lncRNA 39 TCONS-00052400, and it can activate the conversion of long-chain fatty acids. In addition, 40 lncRNA TCONS_00041740 was up-regulated in the high backfat thickness group, and its 41 cis-target gene ACACB was also up-regulated in this group. It has been reported that ACACB is the 42 rate-limiting enzyme in fatty acid oxidation. Conclusions: Since these genes have necessary functions in fat metabolism, the results imply that the lncRNAs detected in our study may affect fat deposition in pigs through regulation of their 45 target genes. In summary, our study explored the regulation of lncRNA and their target genes on 46 fat deposition in pigs and provided new insights for further investigation of the biological 47 functions of lncRNA. In addition, the gene PLA2G12B in the BH group. PLA2G12B A2 group is encoded by this gene and belongs to the phospholipase A2 ( PLA2 ) group of enzymes, which plays a role in lipid hydrolysis by releasing free fatty acids and lysophospholipids [52] . Studies have shown that a reduction of PLA2G12B decreases the amount of serum triglyceride (TG)-rich VLDL particles secreted by the liver, resulting in a reduction in TG content [53] . PLA2G12B can also participate in the pathogenesis of idiopathic membranous nephropathy (iMN) by regulating lipid metabolism [54] . In a previous study by Guan, PLA2G12B -null mice had obvious accumulation of large lipid droplets in the liver, displaying the fatty liver phenotype [55] . These results indicate that the genes exclusively expressed in the high or low backfat groups may also have a certain regulatory effect on lipid metabolism. to and TCONS-00052400 to ACSL3 These results can provide useful information for understanding the regulation of by lncRNA in pigs. However, further genetic experiments are still needed to validate the association of the lncRNA and mRNA functions presented in this study.