Dual-function vector for protein expression in both mammalian cells and Xenopus laevis oocytes
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Dual-function vector for protein expression in both mammalian cells and Xenopus laevis oocytes. / Jespersen, Thomas; Grunnet, M; Angelo, K; Klaerke, D A; Olesen, S P.
I: BioTechniques, Bind 32, Nr. 3, 01.03.2002, s. 536-8, 540.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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T1 - Dual-function vector for protein expression in both mammalian cells and Xenopus laevis oocytes
AU - Jespersen, Thomas
AU - Grunnet, M
AU - Angelo, K
AU - Klaerke, D A
AU - Olesen, S P
PY - 2002/3/1
Y1 - 2002/3/1
N2 - 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.
AB - 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.
KW - Animals
KW - CHO Cells
KW - Cation Transport Proteins
KW - Cricetinae
KW - Electrophysiology
KW - Ether-A-Go-Go Potassium Channels
KW - Gene Expression
KW - Gene Expression Profiling
KW - Genetic Vectors
KW - Kv1.3 Potassium Channel
KW - Mammals
KW - Oocytes
KW - Plasmids
KW - Potassium Channels
KW - Potassium Channels, Voltage-Gated
KW - Transfection
KW - Xenopus laevis
M3 - Journal article
C2 - 11911656
VL - 32
SP - 536-8, 540
JO - BioTechniques
JF - BioTechniques
SN - 0736-6205
IS - 3
ER -
ID: 162549