Analysis and preliminary validation of the molecular mechanism of fat deposition in fatty and lean pigs by high-throughput sequencing.

Fat deposition in muscle includes intramuscular fat (IMF) and intermuscular fat. IMF content is an index of pork quality; however, because IMF content is difficult to measure in vivo in young animals, conventional breeding for IMF content is difficult to carry out. The mechanism and progression of animal fat deposition is not well understood, and there are currently no effective control methods. In this study, using Laiwu and large white pigs as the research subjects and RNA sequencing technology, we analyzed the genetic mechanism of animal fat deposition in pigs. Specifically, we analyzed the features of lncRNAs and their potential target genes. We obtained 464 million clean reads, from which 907 lncRNAs were identified. The cis and trans analysis identified target genes, including genes that were upregulated (286) and downregulated (621) in the fatty and lean pigs. ENSSSCG00000008692_ADD1, ENSSSCG00000023124_ADD1 and ENSSSCG00000005918_DGAT1 were validated as target genes of the lncRNAs and were shown to be closely related to fat deposition. These results provide a basis for studying the different metabolic lncRNA expression of IMF deposition. In addition, as the valuable model animal to study the mechanisms of obesity, pigs may represent a new avenue for studying human obesity.

[Cloning, expression and functional analysis of CuZnSOD gene in swine.].

In order to understand the structure and function of CuZnSOD gene, reveal the effect of the anti-oxidant in swine, and find the molecule marker correlated with meat traits, the cDNA of CuZnSOD gene was cloned and sequenced from muscle of Laiwu black swine by RACE (rapid amplification of cDNA end) techniques. The structure and function of CuZnSOD were analyzed by bioinformatics, and the gene expression profile in different tissues was examined by real-time PCR. The results showed that the full sequence of CuZnSOD cDNA is 658 bp (GU944822), containing 76 bp sequence of 5' UTR and 120 bp sequence of 3' UTR, and coding region (CDS, 462 bp) encodes 153 amino acids. The isoelectric point (pI) of the protein is 6.03, and the molecular weight is 15.9 kDa. There were one O-glycosylation site at the third amino acid and one N-glycosylation site at the eighty-fourth amino acid. The percentage of alpha helix was 1.31%. The alignment similarities of the CDS sequence of swine CuZnSOD with those of cattle, human, rat, and mouse were 87.74%, 87.66%, 83.44%, and 83.23%, and the similarities of amino acid sequence were 90.26%, 94.12%, 92.21%, and 91.50%, respectively. CuZnSOD possesses the typical metal binding ligands (GFHVHQFGDNT). The phylogenic tree based on CuZnSOD protein sequence detected the closest relationship between swine and cattle. CuZnSOD mRNA is a broad-spectrum expression gene, which was detected in brain, heart, spleen, liver, kidney, lung, large intestine, small intestine, spinal cord, muscle, backfat, and stomach. In particular, high expression levels of CuZnSOD mRNA were detected in kidney, small intestine and lung, but low expressions were observed in heart and muscle tissues.

[CDS cloning and relationship between intramuscular fat content and mRNA expression of PID1 gene in pig].

To explore the relationship between the expression of PID1 gene and fat deposition, we cloned CDS of PID1 from porcine fat and muscle tissues by RT-PCR using degenerate primers, and investigated expression of this gene in various tissues (i.e., liver, backfat, and muscle tissues) of different breeds (i.e., Yorkshire, Laiwu, and Lulai Black) by real-time fluorescence quantitative PCR. The results showed that 654 bp CDS of porcine PID1 was obtained by sequencing and was 93.88%, 66.94% and 88.07% identical to those of the human, rat, and Bos taurus, respectively. The expression of PID1 mRNA in various tissues and breeds, on the whole, tended to be liver > fat > muscle and Laiwu > Lulai Black > Yorkshire, respectively. For different breeds, PID1 mRNA abundance in liver had significant difference (P < 0.05), but had no significant differences in fat and muscle tissues between Laiwu and Lulai Black (P > 0.05). For the three groups of Laiwu pigs with high (LWH), intermediate (LWI), and low IMF content (LWL), PID1 mRNA level was higher in liver tissue of LWH than that of LWL significantly (P < 0.05), and was higher in muscle tissue of LWH than that of LWI and LWL significantly (P < 0.05). PID1 mRNA abundance was not correlated with IMF in these three tissues of Laiwu breed, but it was positively correlated with IMF in the tissues of these three breeds (P < 0.05). These results implied that the expression of PID1 may be related to fat deposition.