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Cell-free translation system, method and product

A cell-free, system-based technology, applied in the field of molecular biology, can solve the problems of long time period, low efficiency and high cost, and achieve the effect of enhancing stability and overcoming low efficiency.

Pending Publication Date: 2021-09-17
INSITUTE OF BIOPHYSICS CHINESE ACADEMY OF SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The inventors of the present invention have found that: the main advantage of prokaryotic cell expression is low cost, but many proteins cannot be expressed in prokaryotic cells; eukaryotic cell expression efficiency is high, but there are currently problems of high cost and long time period; Due to the problems of low activity, low efficiency and poor economy in translation technology

Method used

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  • Cell-free translation system, method and product
  • Cell-free translation system, method and product
  • Cell-free translation system, method and product

Examples

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Embodiment 1

[0071] Based on the disclosed embodiments, a cell-free translation system is disclosed, comprising: 5' cap and 3' polyadenylic acid mRNA, amino acid mixture, ATP and mRNA translation mixture.

[0072] The translation system also includes guanosine triphosphate (GTP).

[0073] Preferably, the mRNA structure of the 5' cap and 3' polyadenylic acid is:

[0074] Preferably, the mRNA translation comprises eukaryotic cell lysate supernatant; the amino acid mixture is the published 20 amino acid mixtures.

[0075] This embodiment also discloses a cell-free translation method, comprising the above translation system, including the following steps:

[0076] Step (1) transcribing the mRNA with 5' cap and 3' polyadenylic acid;

[0077] Step (2) adding the mRNA, amino acid mixture, ATP, guanosine triphosphate (GTP) and mRNA into the reaction vessel to react to prepare a mixed solution containing the target protein;

[0078] In step (3), the mixed solution is treated to obtain the target...

Embodiment 2

[0090] Based on the disclosed embodiments, a cell-free translation system is disclosed, comprising: 5' cap and 3' polyadenylic acid mRNA, amino acid mixture, ATP and mRNA translation enzyme source mixture.

[0091] The translation system also includes guanosine triphosphate (GTP).

[0092] Preferably, the mRNA structure of the 5' cap and 3' polyadenylic acid is:

[0093] Preferably, the mRNA translation enzyme source comprises eukaryotic cell lysate supernatant; the amino acid mixture is the disclosed 20 amino acid mixtures.

[0094] This embodiment also discloses a cell-free translation method, comprising the above translation system, including the following steps:

[0095] Step (1) transcribing the mRNA with 5' cap and 3' polyadenylic acid;

[0096] Step (2) Add the mRNA, amino acid mixture, ATP, guanosine triphosphate (GTP) and mRNA translation enzyme source into the reaction vessel, and react to prepare a mixed solution containing the target protein;

[0097] In step (3...

Embodiment 3

[0109] Based on the disclosed embodiments, a cell-free translation system is disclosed, comprising: 5' cap and 3' polyadenylic acid mRNA, amino acid mixture, ATP and mRNA translation enzyme source mixture.

[0110] The translation system also includes guanosine triphosphate (GTP).

[0111] Preferably, the mRNA structure of the 5' cap and 3' polyadenylic acid is:

[0112] Preferably, the mRNA translation enzyme source comprises eukaryotic cell lysate supernatant; the amino acid mixture is a mixture of 20 amino acids.

[0113] This embodiment also discloses a cell-free translation method, comprising the above translation system, including the following steps:

[0114] Step (1) transcribing the mRNA with 5' cap and 3' polyadenylic acid;

[0115] Step (2) Add the mRNA, amino acid mixture, ATP, guanosine triphosphate (GTP) and mRNA translation enzyme source into the reaction vessel, and react to prepare a mixed solution containing the target protein;

[0116] In step (3), the mi...

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Abstract

The invention relates to the field of molecular biology, in particular to a cell-free translation system, method and product. The translation system comprises mRNA of 5'cap and 3 'polyadenylic acid, an amino acid mixture, an energy molecule and a mixture of mRNA translation enzyme sources; the energy molecule comprises phosphocreatine, ATP (adenosine triphosphate) and guanosine triphosphate (GTP). Through mRNA transcription, in-vitro transcription and gel electrophoresis analysis, the translation system can rapidly obtain a large amount of target proteins and has the following characteristics: an extracellular translation technology is adopted to overcome the problem that a prokaryotic cell expression part gene cannot be expressed; the problems of long expression cycle and low efficiency of eukaryotic cells are solved by adopting an extracellular translation technology; the problems of low activity and poor efficiency of the original extracellular translation technology are solved by adopting a strategy of adding energy micromolecules such as phosphocreatine; and the stability, the receptor binding speed and the overall translation speed are enhanced by adopting mRNA of 5 ' cap and 3' polyadenylic acid.

Description

technical field [0001] The technology belongs to the field of molecular biology, and in particular relates to a cell-free translation system, method and product. Background technique [0002] The final product of gene expression mainly exists in the form of protein except for non-coding RNA. The existing protein expression technologies mainly include expression in prokaryotic bacteria, expression in eukaryotic insect cells, and expression in human engineered cells. The inventors of the present invention have found that: the main advantage of prokaryotic cell expression is low cost, but many proteins cannot be expressed in prokaryotic cells; eukaryotic cell expression efficiency is high, but there are currently problems of high cost and long time period; Translation technology has problems such as low activity, low efficiency, and poor economy. Contents of the invention [0003] The researchers of the present invention have found through research that the "cap" structure a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G16B20/20
CPCG16B20/20
Inventor 张晓荣赵谦
Owner INSITUTE OF BIOPHYSICS CHINESE ACADEMY OF SCIENCES
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