Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-Shan, Taoyuan 333, Taiwan a Department of Medical Biotechnology and Laboratory Science, College of Medicine, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-Shan, Taoyuan 333, TaiwanDepartment of Ophthalmology, Chang Gung Memorial Hospital,Department of Medicine, College of Medicine, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-Shan, Taoyuan 333, TaiwanFu-Hsing Street, Kwei-Shan, Taoyuan 333, TaiwanDepartment of Safety, Health and Environmental Engineering, Ming Chi University of Technology, 84 Gungjuan Rd., Taishan Dist, New Taipei City 243, Taiwan g Department of Chemical and Materials Engineering, College of Engineering, Chang Gung University, 259 Wen-Hwa 1st Road, Kwei-Shan, Taoyuan 333, Taiwanf Department of Chemical Engineering, Ming Chi University of Technology, 84 Gungjuan Rd., Taishan Dist, New Taipei City 243, Taiwan
Background Internal ribosome entry sites allow the translation of a transcript independent of its cap structure. They are distributed in some viruses and cellular RNA. The element is applied in dual gene expression in a single vector. Although it appears the lower efficiency of IRES-mediated translation than that of cap-dependent translation, it is with the crucial needs to know the precise differences in translational efficacy between upstream cistrons and downstream cistrons before applyin...更多
Background Internal ribosome entry sites allow the translation of a transcript independent of its cap structure. They are distributed in some viruses and cellular RNA. The element is applied in dual gene expression in a single vector. Although it appears the lower efficiency of IRES-mediated translation than that of cap-dependent translation, it is with the crucial needs to know the precise differences in translational efficacy between upstream cistrons and downstream cistrons before applying the bicistronic vector in biomedical applications. Material and Methods This study aimed to provide real examples and showed the precise differences for translational efficiency dependent upon target gene locations. We generated various bicistronic constructs with quantifiable reporter genes as upstream and downstream cistrons of the encephalomyocarditis virus IRES to precisely evaluate the efficacy of IRES-mediated translation in mammalian cells. Results There was no significant difference in protein production when the reporter gene was cloned as an upstream cistron. However, lower levels of protein production were obtained when the reporter gene was located downstream of the IRES. Moreover, in the presence of an upstream cistron, a markedly reduced level of protein production was observed. Conclusions Our findings demonstrate the version of the EMCV IRES that is provided in many commercial vectors is relatively less efficient than cap-dependent translation and provide valuable information regarding the utilization of IRES to facilitate the expression of more than one protein from a transcript.收起
