Abstract
Mammalian expression vectors are used to overexpress genes of interest in mammalian cells. High temperature requirement protein A1 (HtrA1), used as a specific target, was expressed from the pHA-M-HtrA1 plasmid in HEK293T cells, inducing cell death. Expression of HtrA1 was driven by the pHA-M-HtrA1 mammalian expression vector in E. coli resulting in growth suppression of E. coli in an HtrA1 serine protease-dependent manner. By using various combinations of promoters, target genes and N-terminal tags, the T7 promoter and N-terminal HA tag in the mammalian expression vector were shown to be responsible for expression of target genes in E. coli. Thus the pHA-M-HtrA1 plasmid can be used as a novel, rapid pre-test system for expression and cytotoxicity of the specific target gene in E. coli before assessing its functions in mammalian cells.
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Acknowledgments
This stduy was supported by the National Nuclear R&D Program through the National Research Foundation (NRF) of Korea, which is funded by the Ministry of Education, Science and Technology (MEST) (2011-0018783); and NRF Grant funded by the Korean Government MEST (2010-0029422); and a Grant of the Korea Health Technology R&D Project, Ministry of Health and Welfare, Republic of Korea (A101099).
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J.-M. Moon and G.-Y. Kim contributed equally to this study.
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Moon, JM., Kim, GY. & Rhim, H. A new idea for simple and rapid monitoring of gene expression: requirement of nucleotide sequences encoding an N-terminal HA tag in the T7 promoter-driven expression in E. coli . Biotechnol Lett 34, 1841–1846 (2012). https://doi.org/10.1007/s10529-012-0966-8
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DOI: https://doi.org/10.1007/s10529-012-0966-8