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In-vitro Mutagenesis

In-vitro Mutagenesis

To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes.

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Updated: May 23, 2025

Preparation of Cell-lines for Conditional Knockdown of Gene Expression and Measurement of the Knockdown Effects on E4orf4-Induced Cell Death

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Published on: October 21, 2012

通过有针对性的整合来评估CHO培养表现的异位基因效率.

David Catalán-Tatjer¹, Saravana Kumar Ganesan¹, Iván Martínez-Monje¹

¹The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Lyngby 2800, Denmark.

ACS synthetic biology

|April 23, 2025

概括

过度表达中国仓鼠卵巢 (CHO) bcl-2显著改善了生物制药生产. 这项研究建立了一个有针对性的基因整合方法,以克服细胞培养性能代谢工程中的挑战.

关键词:

在CHO细胞中.这是一种反apoptosis.有针对性的整合.

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Last Updated: May 23, 2025

Preparation of Cell-lines for Conditional Knockdown of Gene Expression and Measurement of the Knockdown Effects on E4orf4-Induced Cell Death

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科学领域:

生物技术是生物技术.
细胞生物学细胞生物学
生物加工是一种生物加工.

背景情况:

中国仓鼠卵巢 (CHO) 细胞对于生物制药生产至关重要,但细胞死亡限制了培养性能.
细胞亡 (编程细胞死亡) 是影响CHO细胞培养时间和密度的关键因素.
之前的研究由于随机基因整合的克隆变异而面临挑战.

研究的目的:

开发同源性CHO细胞系,用于研究抗瘤基因.
为了确定增强生物制药生产的特定抗菌基因.
建立一种可靠的方法来测试CHO细胞工程中的候选基因.

主要方法:

利用复合酶介导的磁带交换来创建表达 erythropoietin 和各种抗apoptotic 基因 (bcl-2, bcl-xL, mcl-1, bhrf-1) 的同源性 CHO 细胞系.
在用甲酸盐的批量培养和使用ambr15微生物反应器系统在食批量中测试细胞系.
采用多重化定量蛋白质组学来描述代谢差异.

主要成果:

过度表达CHO来源的bcl-2显著提高了生产力.
观察到的表型变异取决于过度表达的特定抗瘤基因.
成功建立了针对性基因测试的方法.

结论:

针对性地整合抗apoptotic 基因,特别是 CHO bcl-2,可以提高 CHO 细胞培养的生产力.
开发的方法使得更多可比的代谢工程策略成为可能.
这项研究克服了与CHO细胞系发育中的克隆变异相关的先前挑战.