Irisin recombinant protein and synthetic method thereof
A recombinant protein and protein technology, which is applied in the field of genetic engineering, can solve the problems such as the localization and expression analysis and identification of Irisin protein that cannot be used, and achieve the effect of accurate research and localization analysis.
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Embodiment 1
[0052] An Irisin recombinant protein, the N-terminal of the Irisin recombinant protein has a His-tag.
Embodiment 2
[0054] A kind of synthetic method of the DNA sequence of Irisin recombinant protein, comprises the following steps:
[0055] (1) Design and sequence acquisition of Irisin prokaryotic expression plasmid.
[0056] (2) Obtain the cohesive Irisin sequence.
[0057] (3) Obtain the pET28a(+) linearly linked plasmid with cohesive ends.
[0058] (4) Ligate the pET28a(+) plasmid and the Irisin sequence product.
Embodiment 3
[0060] The design and sequence acquisition of Irisin prokaryotic expression plasmids include the following steps:
[0061] (1) Design of sequences and primers:
[0062] A. Check the FNDC5 amino acid sequence and FNDC5 cDNA sequence from NCBI and Ensembl websites. The FNDC5 amino acid sequence includes the Irisin amino acid sequence, and the FNDC5 cDNA sequence includes the Irisin cDNA sequence.
[0063] The sequence of Irisin is highly conserved, and the amino acid sequences of human, rat, and mouse are completely consistent. In the present invention, the sequence of mouse is selected for recombination and synthesis, and the mRNA containing FNDC5 sequence obtained from the mRNA of mouse preadipocyte strain 3T3-L1 full sequence.
[0064] B. Design primers according to NdeI and XhoI enzyme digestion. The 5' end of the primer has a sequence of enzyme cutting points, and the rest are sequences located at both ends of Irisin. The codons of amino acids after connection are consiste...
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