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Vector pGEMT/rhPA, construction method of vector and application of vector in preparation method of efficient recombinant human plasminogen activator

A carrier, rhpa technology, which is applied in recombinant DNA technology, biochemical equipment and methods, and the introduction of foreign genetic material using a carrier, can solve the problems of high clinical treatment costs, increased production costs, and prone to adverse reactions.

Active Publication Date: 2014-02-12
YANGZHOU UNIV
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Problems solved by technology

As a result, production costs increase, resulting in expensive clinical treatment costs
In addition, prokaryotic products have poor compatibility with the human body and are prone to adverse reactions, which also reduces the safety of the drug and increases the risk of clinical application

Method used

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  • Vector pGEMT/rhPA, construction method of vector and application of vector in preparation method of efficient recombinant human plasminogen activator
  • Vector pGEMT/rhPA, construction method of vector and application of vector in preparation method of efficient recombinant human plasminogen activator
  • Vector pGEMT/rhPA, construction method of vector and application of vector in preparation method of efficient recombinant human plasminogen activator

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Experimental program
Comparison scheme
Effect test

Embodiment Construction

[0421] 1. Carrier construction:

[0422] The technical route of carrier construction is attached figure 1 , the operating steps for constructing the mammary gland characteristic expression component are as follows:

[0423] (1) Materials

[0424] 1.1 Experimental materials and instruments

[0425] DNA amplification instrument (S1000, BIO-RAD, USA), DNA electrophoresis instrument, desktop refrigerated high-speed centrifuge, small desktop high-speed centrifuge (Eppendorf, Germany), constant temperature shaking incubator, redistilled water device, constant temperature water bath shaker, Water-proof electric heating constant temperature incubator, digital gel imager (GIS-1000, Shanghai Tianneng Technology Co., Ltd.), upper dish electronic balance (FA1604, Shanghai Precision Balance), PH detector (HA405-K2, Mettlerto Lido Shanghai Company), ice machine (SIM-F124 Japan Sanyo Company), ultra-clean bench (Suzhou Purification Instrument Equipment Factory), vacuum dryer (Speed ​​Va...

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Abstract

The invention provides a vector pGEMT / rhPA, a construction method of the vector and an application of the vector in a preparation method of an efficient recombinant human plasminogen activator, and belongs to the genetic and cell engineering technologies in the bioengineering field. The construction method of the vector pGEMT / rhPA comprises the following steps of firstly constructing a vector pGEMT / 5', secondly constructing a vector pGEMT / 5'+3'P, and finally constructing a vector pGEMT / rhPA, wherein the vector pGEMT / rhPA HAS 1068bp at a position of 9203-10269, and encodes 356 pieces of amino acid. Based on experiments, the recombinant product expressed by the vector pGEMT / rhPA has the advantages of high content and high specific activity; the vector pGEMT / rhPA can be applied to development of human thrombolytic agents.

Description

technical field [0001] The invention belongs to the genetic engineering and cell engineering technology in the field of bioengineering, in particular to the technical field of large-scale recombinant human plasminogen activator (rhPA) production. Background technique [0002] Animal mammary gland bioreactor technology refers to the use of mammalian mammary gland-specific promoter elements to construct transgenic animals, directing the expression of foreign genes in the mammary glands of transgenic animals, and obtaining recombinant proteins from animal milk, which has high commercial value. Value. Rabbits have the advantages of high lactation, short gestation period, and large number of litters per litter. So far, the application of rabbit mammary gland bioreactor to produce recombinant human tissue plasminogen activator has not been reported. [0003] Thrombotic disease is one of the major diseases that endanger human life and health. At present, natural human tissue plas...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/63C12N15/66C12N15/85C12N9/72
Inventor 成勇成姚斌宋绍征葛欣
Owner YANGZHOU UNIV
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