Method for preparing aspartate aminotransferase and application of aspartate aminotransferase

An aspartate amino, transferase technology, applied in the fields of genetic engineering and protein expression, can solve the problems of lack of experience and strength in diagnostic reagents, few domestic products, etc., and achieves high finished product yield, low preparation cost, and simple operation. Effect

Inactive Publication Date: 2012-12-05
BEIJING XIANGDA SHENGJI SCI & TECH CONSULTING
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Problems solved by technology

Almost none of the major domestic manufacturers of diagnostic reagents has the experience and strength to produce...

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  • Method for preparing aspartate aminotransferase and application of aspartate aminotransferase
  • Method for preparing aspartate aminotransferase and application of aspartate aminotransferase

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Embodiment Construction

[0047]The present invention will now be described more fully with reference to the accompanying drawings, in which exemplary embodiments of the invention are shown. This invention may, however, be embodied in many different forms and should not be construed as limited to the exemplary embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art.

[0048] The invention establishes a method for preparing aspartate aminotransferase with simple operation, rapid operation and low cost. The principle of this method mainly lies in: inserting the optimized aspartate aminotransferase coding gene into the constructed expression vector, and inducing its high-level expression in Escherichia coli, the method is specifically as follows:

[0049] 1. Construction of aspartate aminotransferase expression vector:

[0050] Transform the coding gene sequenc...

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Abstract

The invention discloses a method for preparing aspartate aminotransferase and application of the aspartate aminotransferase. The method includes: modifying the coding gene sequence of humanized aspartate aminotransferase subjected to gene synthesis, and removing escherichia coli rare codons; inserting the modified aspartate aminotransferase coding gene sequence into an expression vector, and using an escherichia coli strain to transform expression; inoculating strain for transforming expression to LB (Luria-Bertani) medium for IPTG (isopropyl-beta-d-thiogalactoside) induction; and homogenizing and centrifuging obtained bacteria, and enabling the centrifuged bacteria to pass through the Ni-NTA column for purification. The method for preparing the aspartate aminotransferase is extremely simple in operation, low in preparation cost and high in finished product yield and purity; the aspartate aminotransferase has high biological activity and can be used for animal immunization and screening related antibodies by serving as antigen substances; and the method and the aspartate aminotransferase lay the foundation for establishing a badly-needed calibrator preparation platform as early as possible for domestic clinical chemistry diagnosis and developing diagnostic reagent standard substances and references.

Description

technical field [0001] The invention belongs to the field of genetic engineering and protein expression, and relates to a method for efficiently preparing aspartate aminotransferase using bioengineering technology. The product can be used as an antigenic substance for animal immunization, to screen related antibodies, and to calibrate for clinical chemical diagnosis. It lays the foundation for the preparation and development of products, standard materials and reference products. Background technique [0002] Aspartate aminotransferase (aspartate aminotransferase AST), formerly known as aspartate aminotransferase (glutamic oxaloacetic transaminase GOT), is an important transaminase in the body and widely exists in various tissues in the body. AST has the highest activity in the heart. The content in normal human serum is very small. Under normal circumstances, the range of aspartate aminotransferase from liver cells in the blood is 0-40U / L (unit). [0003] When cardiomyocy...

Claims

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Application Information

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IPC IPC(8): C12N15/54C12N15/70C12N9/10C12Q1/52G01N33/573
Inventor 肖飞黄薇王大光
Owner BEIJING XIANGDA SHENGJI SCI & TECH CONSULTING
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