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Cat serum albumin recombinant protein and efficient expression method thereof in pichia pastoris

A kind of serum albumin, recombinant protein technology, applied in the field of cat serum albumin recombinant protein and its high expression in Pichia pastoris

Active Publication Date: 2020-05-08
泰州博莱得利生物科技有限公司 +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In addition, due to the influence of cat serum raw materials, there are no cat serum albumin-related products on the market at home and abroad

Method used

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  • Cat serum albumin recombinant protein and efficient expression method thereof in pichia pastoris
  • Cat serum albumin recombinant protein and efficient expression method thereof in pichia pastoris
  • Cat serum albumin recombinant protein and efficient expression method thereof in pichia pastoris

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Experimental program
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Effect test

Embodiment

[0049] 1. Synthesis of cat serum albumin gene: based on the gene sequence of wild-type cat serum albumin (as shown in SEQ ID NO: 1 and figure 1 Shown), (amino acid sequence is shown in SEQ ID NO: 2 and figure 2 shown), the DNA coding sequence of feline serum albumin was optimized with codon optimization software (http: / / www.jcat.de) to obtain the DNA sequence (such as SEQ ID NO : 3 shown and image 3 shown), the complete sequence was synthesized by Shanghai Sangon. The amino acid sequence of the feline serum albumin recombinant protein deduced by it is shown in SEQ ID No.4 and Figure 4 shown.

[0050] 2. Construction and identification of the recombinant plasmid: combine the coding sequence of the optimized feline serum white recombinant protein with the Pichia pastoris expression vector pPIC9K (such as Figure 5 shown) were digested with SnaB I and Not I respectively.

[0051] Figure 5 In , the DNA sequence encoding feline serum albumin is inserted between the SnaB I...

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Abstract

The present invention provides a cat serum albumin recombinant protein and an efficient expression method thereof in pichia pastoris, and relates to a recombinant nucleotide sequence of the cat serumalbumin (as shown in SEQ ID NO:3), a vector containing the nucleotide sequence, and an engineered cell containing the aforementioned vector. Based on a natural sequence of a cat serum albumin gene, codons are optimized to enable the gene to be more suitable to be expressed in yeast cells, the modified encoded cat serum albumin DNA sequence is cloned into a pichia pastoris expression vector pPICK9,a pichia pastoris expression strain GS115 is transformed, and G418 pressure screening is conducted to obtain a clone train that efficiently expresses the cat serum albumin with an expression level upto 2 g / L. The protein expressed by the method is close to natural, high in expression efficiency, simple in operation process and suitable for large-scale industrial production.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to feline serum albumin recombinant protein and its high-efficiency expression method in Pichia pastoris. Background technique [0002] Feline serum albumin (Feline serum albumin, FSA) is a single-chain protein composed of 584 amino acids, with a molecular mass of 65.85kDa. It is a protein with the highest content and more functions in serum. The maintenance of plasma colloid osmotic pressure mainly depends on albumin in plasma (serum albumin constitutes about 75% to 80% of plasma colloid osmotic pressure). The osmotic pressure produced by 1 g of albumin is equivalent to the hemodynamic effect obtained by 20 ml of plasma or 40 ml of whole blood. [0003] Feline albumin is synthesized by liver cells. When liver disease occurs, the synthesis of albumin will be impaired, especially when liver cirrhosis occurs, the synthesis of albumin will be significantly reduced, leading to...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/12C12N15/81C12N1/19C07K14/765C12R1/84
CPCC07K14/765C12N15/815
Inventor 刘继红纪志辉石文达刘方董昊王宏伟
Owner 泰州博莱得利生物科技有限公司
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