首页> 外国专利> A Cell Model For In Vitro Evaluation Of Compound-Induced Skin Sensitization And A Constructing Method Therefor

A Cell Model For In Vitro Evaluation Of Compound-Induced Skin Sensitization And A Constructing Method Therefor

机译：复合诱导的皮肤致敏性体外评估的细胞模型及其构建方法

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摘要

A cell model for in vitro evaluation of compound-induced skin sensitization and a constructing method therefor. The method for constructing the cell model comprises the steps of: designing and constructing an sgRNA expression vector based on CRISPR/Cas9 vector system; designing and constructing a homologous recombinant vector capable of knocking a reporter gene linked to a self-cleaving peptide sequence into a specific site of the expression frame of the HMOX1 gene; co-transfecting the homologous recombinant vector, an hCas9 plasmid and the sgRNA expression vector into a cell, and carrying out monoclonal expansion to obtain the cell model. The present invention obtains a HaCaT cell model in which a luciferase gene is knocked in before the stop codon of the HMOX1 gene by combination of CRISPR/CAS9 and a monoclonal cell technique. The cell model realizes synchronous expression of the luciferase gene and the HMOX1 gene, thereby effectively distinguishing sensitizing compounds from non-sensitizing compounds.

机译：用于体外评估化合物诱导的皮肤致敏作用的细胞模型及其构建方法。构建细胞模型的方法包括以下步骤：基于CRISPR / Cas9载体系统设计和构建sgRNA表达载体。设计并构建能够将与自切割肽序列连接的报道基因敲入HMOX1基因表达框架的特定位点的同源重组载体;将同源重组载体，hCas9质粒和sgRNA表达载体共转染到细胞中，进行单克隆扩增，得到细胞模型。本发明获得了一种HaCaT细胞模型，其中通过结合CRISPR / CAS9和单克隆细胞技术，在HMOX1基因的终止密码子之前敲入了荧光素酶基因。该细胞模型实现了荧光素酶基因和HMOX1基因的同步表达，从而有效地区分了敏化化合物和非敏化化合物。

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公开/公告号US2020140856A1

专利类型
公开/公告日2020-05-07

原文格式PDF
申请/专利权人 SOUTH CHINA UNIVERSITY OF TECHNOLOGY;
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申请/专利号US201816611818
发明设计人 LIZHEN HUANG;GUORUI ZHONG;HAOJIAN LI;SHUILIN XIE;SHIHUI PANG;JING BAI;CHANGSHENG HE;XINYI DU;QIXIAO ZHANG;HAIJIE WANG;HONGLI DU;
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申请日2018-10-29
分类号C12N15/11;C12N9/22;C12N15/85;C12N5/071;C12N5/0784;C12N5/0786;
国家 US
入库时间 2022-08-21 11:20:02

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