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Enhanced detoxification function gene fragment and modified hepg2 cells

A technology of detoxification function and gene fragments, applied in the field of genetic engineering, to achieve the effect of improving detoxification ability

Active Publication Date: 2022-06-21
WUHAN TOGO MEDITECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The purpose of the present invention is to overcome the defects of existing HepG2 cells, and provide a HepG2 cell that enhances detoxification function gene fragments and transformation

Method used

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  • Enhanced detoxification function gene fragment and modified hepg2 cells
  • Enhanced detoxification function gene fragment and modified hepg2 cells
  • Enhanced detoxification function gene fragment and modified hepg2 cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0031] Embodiment: the scheme of HepG2 cell line transformation:

[0032] 1) The entire genome was assembled into pTBSCH2-OTC-ARG1-OATP1B1-Bst, and linearized. The sequence is shown in SEQNo.2.

[0033] 2) Digest HepG2 cells (ATCC, American Type Culture Collection Center) that night, and its sequence can be found in (NCBIGenBank), and plant them in T25 at a density of 40%-50%, and after overnight attachment, the next morning Perform transfection.

[0034] 3) Transfection reagents and materials

[0035]

[0036] (plasmid:transposon=10:1)

[0037] Mix well, let it stand for 5min, add it to T25, and change the medium according to the state of the cells.

[0038] 4) Five days after the cells were transfected, they were digested and counted, and about 1000-2000 cells per well were seeded into a 96-well plate.

[0039] 5) After the cells have grown to a certain number, continue to seed the plate for screening. After the preliminary detection of negative cells (HepG2), screen...

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Abstract

The invention discloses a gene fragment for enhancing detoxification function and a transformed HepG2 cell, wherein the gene fragment for enhancing detoxification function is composed of three gene fragments of OTC, ARG1 and OATP1B1 in sequence. In HepG2 cells transformed with this gene fragment, the expression of albumin was doubled; the expression of aspartate aminotransferase was increased by 10%; the secretion of key proteins related to endogenous detoxification ability in the cytochrome P450 family was increased, and the detoxification ability was improved: the secretion of CYP2C9 was increased by 5 times, CYP2D6 secretion increased by 50 times, CYP2A6 secretion increased by 2 times, and CYP3A4 secretion increased by 100 times.

Description

technical field [0001] The invention relates to the field of genetic engineering, in particular to a gene fragment for enhancing detoxification function and transformed HepG2 cells. Background technique [0002] HepG2 cells are a type of liver cancer cells. The cells secrete a variety of plasma proteins: albumin, α2-macroglobulin, plasminogen, transferrin, etc. [0003] Existing HepG2 cells have blood ammonia metabolism and detoxification defects when they are used for blood purification. Contents of the invention [0004] The purpose of the present invention is to overcome the defects of the existing HepG2 cells, and provide a gene segment for enhancing detoxification function and the transformed HepG2 cells. [0005] In order to achieve the above purpose, the present invention firstly provides a gene segment for enhancing detoxification function, which is composed of three gene segments of OTC, ARG1, and OATP1B1 in sequence, wherein the secreted proteins corresponding ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/62C12N15/85C12N15/65C12N5/10C12R1/91
CPCC12N9/1018C12N9/78C12N15/85C12N15/65C07K14/47C12Y201/03003C12Y305/03001C07K2319/00
Inventor 王柯丰明乾望威
Owner WUHAN TOGO MEDITECH CO LTD
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