An Internal Ribosome Entry Site (IRES) Mutant Library for Tuning Expression Level of Multiple Genes in Mammalian Cells
An Internal Ribosome Entry Site (IRES) Mutant Library for Tuning Expression Level of Multiple Genes in Mammalian Cells
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DOI：
10.1371/journal.pone.0082100
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发表时间：
2013-12-09
期刊：
影响因子：
3.7
通讯作者：
Yang, Yuansheng
中科院分区：
文献类型：
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作者：
Koh, Esther Y. C.;Ho, Steven C. L.;Yang, Yuansheng
通过将8月10日、11日和12日的翻译起始密码子突变为非8月的三联体，产生了一组强度不同的突变的脑心肌炎病毒(EMCV)内部核糖体进入位点(IRES)元件。它们能够在瞬时和稳定的转染中在哺乳动物细胞中广泛地控制多个基因的相对表达。在不同的哺乳动物细胞系中，每个IRES突变体的相对强度保持相似，并且不是基因特异性的。表达的蛋白质具有正确的分子量。这些IRES突变体优化了重链上的轻链表达，提高了单抗的表达水平和稳定表达的质量。这组IRES突变体的用途可以扩展到其他应用，如合成生物学、研究蛋白质及其复合体之间的相互作用、细胞工程、多亚单位蛋白质生产、基因治疗以及将体细胞重新编程为干细胞。
A set of mutated Encephalomyocarditis virus (EMCV) internal ribosome entry site (IRES) elements with varying strengths is generated by mutating the translation initiation codons of 10th, 11th, and 12th AUG to non-AUG triplets. They are able to control the relative expression of multiple genes over a wide range in mammalian cells in both transient and stable transfections. The relative strength of each IRES mutant remains similar in different mammalian cell lines and is not gene specific. The expressed proteins have correct molecular weights. Optimization of light chain over heavy chain expression by these IRES mutants enhances monoclonal antibody expression level and quality in stable transfections. Uses of this set of IRES mutants can be extended to other applications such as synthetic biology, investigating interactions between proteins and its complexes, cell engineering, multi-subunit protein production, gene therapy, and reprogramming of somatic cells into stem cells.