Chinese Journal of Biotechnology 2002;18(3):308-312

Construction of eukaryotic expression vector using neomycin-resistance gene mutant as selectable marker.

Chuan GAO 1 2 ; Xu-Dong ZHU ; Xiao-Wei ZHOU ; Fang YU ; Bai-Song LU ; Pei-Tang HUANG

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Country

China

Language

Chinese

Abstract

Neomycin-resistance gene is widely used as a selectable marker in eukaryotic expression vector. It codes neomycin phosphotransferase II (NPT II) which confers resistance to various aminoglycoside antibiotic such as G418 and kanamycine. In this work, by site-directed mutagenesis the neo gene mutant was obtained. The expression vector pmDNA using the neo gene mutant as selectable marker has been constructed. After inserting interest luciferase gene, the expression plasmid pmDNAluc + was stably transfected CHO-K1 cells. As a result, the expression positive ratio reaches to approximate 95% and the ratio of high expression colonies is apparently higher than the controls.