Dual-function vector for protein expression in both mammalian cells and Xenopus laevis oocytes.
Both Xenopus laevis oocytes and mammalian cells are widely used for heterologous expression of several classes of proteins, and membrane proteins especially, such as ion channels or receptors, have been extensively investigated in both cell types. A full characterization of a specific protein will often engage both oocytes and mammalian cells. Efficient expression of a protein in both systems have thus far only been possible by subcloning the cDNA into two different vectors because several different molecular requirements should be fulfilled to obtain a high protein level in both mammalian cells and oocytes. To address this problem, we have constructed a plasmid vector, pXOOM, that can function as a template for expression in both oocytes and mammalian cells. By including all the necessary RNA stability elements for oocyte expression in a standard mammalian expression vector, we have obtained a dual-function vector capable of supporting protein production in both Xenopus oocytes and CHO-K1 cells at an expression level equivalent to the levels obtained with vectors optimized for either oocyte or mammalian expression. Our functional studies have been performed with hERGI, KCNQ4, and Kv1.3 potassium channels.
Jespersen
T
T
University of Copenhagen, Department of Medical Physiology, The Panum Institute, Denmark. tjespersen@mfi.ku.dk
Grunnet
M
M
Angelo
K
K
Klaerke
D A
DA
Olesen
S P
SP
D000818:Animals; D016466:CHO Cells; D027682:Cation Transport Proteins; D006224:Cricetinae; D004594:Electrophysiology; D051638:Ether-A-Go-Go Potassium Channels; D015870:Gene Expression; D020869:Gene Expression Profiling / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D051664:Kv1.3 Potassium Channel; D008322:Mammals; D009865:Oocytes; D010957:Plasmids / Q000235:genetics*; D015221:Potassium Channels / Q000235:genetics; D024642:Potassium Channels, Voltage-Gated; D014162:Transfection; D014982:Xenopus laevis / Q000235:genetics*
D027682:Cation Transport Proteins; D051638:Ether-A-Go-Go Potassium Channels; C092509:KCNH6 protein, human; D051664:Kv1.3 Potassium Channel; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated