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Development of the Mammalian Expression Vector System that can be Induced by IPTG and/or Lactose

Journal of microbiology and biotechnology, v.30 no.8, 2020년, pp.1124 - 1131  

Myung, Seung-Hyun (Department of Biochemistry and Molecular Biology, Chosun University School of Medicine) ,  Park, Junghee (Department of Biochemistry and Molecular Biology, Chosun University School of Medicine) ,  Han, Ji-Hye (Department of Biochemistry and Molecular Biology, Chosun University School of Medicine) ,  Kim, Tae-Hyoung (Department of Biochemistry and Molecular Biology, Chosun University School of Medicine)

Abstract AI-Helper 아이콘AI-Helper

Techniques used for the regulation of gene expression facilitate studies of gene function and treatment of diseases via gene therapy. Many tools have been developed for the regulation of gene expression in mammalian cells. The Lac operon system induced with isopropyl β-D-1-thiogalactopyranosi...

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제안 방법

  • Hence, we thought that the five operators massively block the binding of RNA polymerase II to CMV promoter-mini region through the LacI repressors, and significantly reduce the expression of GFP after inducer treatment. Thus, we constructed two modified inducible vectors by deleting the Lac operators from pCalo5-GFP-LacI vector to improve the expression level of our inducible vector system (Figs. 5A and B). We termed the resulting two vectors as pCalo5-GFP-LacI-Del-II and pCalo5-GFP-LacI-Del-III.

이론/모형

  • 1% sodium dodecyl sulfate [SDS], 150 mM sodium chloride [NaCl], 2 mM ethylenediaminetetraacetic acid [EDTA], and 1/1000 protease inhibitor cocktail, Sigma-Aldrich). Protein concentrations were quantified with the bicinchoninic acid (BCA) assay method (ThermoFisher Scientific, USA). About 30 μg of proteins were separated with SDS polyacrylamide gel electrophoresis and the separated protein bands were transferred onto polyvinylidene fluoride (PVDF) membranes (GE Healthcare Life Science, USA) using an electro-transfer machine at 100 mA for 400 min.
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참고문헌 (30)

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