Cloning, prokaryotic expression, purification, and functional verification of the insulin gene in black carp (Mylopharyngodon piceus)
2023
Zhang, Jiahua | Shen, Yubang | Dai, Yafan | Xu, Xiaoyan | Su, Yuhong | Li, Jiale
In this paper, we studied the structure, expression and function of black carp insulin gene. The complete Mylopharyngodon piceus insulin (Mp-Ins) gene is 1,965 bp long and includes a 1,499 bp 5ʹ-untranslated region (UTR), a 139 bp 3′-UTR with a poly(A) tail, and an open reading frame (ORF) of 327 bp. The predicted molecular weight of the recombinant Mp-Ins(rMp-Ins) protein is 11.87 kDa. The mRNA expression of Mp-Ins is upregulated in the brain and liver. After the injection of rMp-Ins, Mp-Ins mRNA transcript abundance was significantly upregulated in the liver. The rMp-Ins protein could inhibit the concentration of glycogen phosphorylase (GP), growth hormone (GH), fatty acid synthase (FAS), and insulin-like growth factors-1 (IGF-1), and it also significantly increased the concentration of PI3K. Additionally, the injection of rMp-Ins did not have a significant impact on the glucose-6-phosphatase (G6Pase) content in blood. In situ hybridization results showed that the positive signal of the Mp-Ins gene was mainly concentrated in the cell nucleus of brain tissue and the cell membrane of liver tissue and muscle tissue. Together, these results demonstrated that Mp-Ins plays an important role in growth and metabolism in M. piceus.
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