High efficiency transfection of glioma cell lines and primary cells for overexpression and RNAi experiments.
In order to investigate the impact of signalling proteins on the phenotype and malignant behavior of glioblastoma cells, we optimized the transfection procedure of human glioblastoma cell lines U251, U373, GaMG and of primary cells obtained from a patient's tumor using nucleofection technology in conjunction with plasmid pmaxGFP. We describe the optimization procedure, show that a high percentage of the cells can be transfected and that nucleofection does not cause phenotypic alterations of the cells. Therefore, we conclude that nucleofection is a highly efficient tool to deliver plasmids for transient protein overexpression and siRNA for specific protein knock-down to different glioblastoma cell lines or primary cells.
Hagemann
Carsten
C
Department of Neurosurgery, Tumorbiology Laboratory, University of Würzburg, Josef-Schneider-Str. 11, D-97080 Würzburg, Germany. hagemann_c@klinik-uni-wuerzburg.de
Meyer
Christoph
C
Stojic
Jelena
J
Eicker
Sven
S
Gerngras
Stefanie
S
Kühnel
Siglinde
S
Roosen
Klaus
K
Vince
Giles Hamilton
GH
D015153:Blotting, Western; D001932:Brain Neoplasms / Q000235:genetics* / Q000378:metabolism*; D045744:Cell Line, Tumor; D002465:Cell Movement; D002467:Cell Nucleus / Q000235:genetics; D049109:Cell Proliferation; D003584:Cytological Techniques; D018274:Electroporation; D005904:Glial Fibrillary Acidic Protein / Q000096:biosynthesis / Q000235:genetics; D005910:Glioma / Q000235:genetics* / Q000378:metabolism*; D049452:Green Fluorescent Proteins; D006801:Humans; D007150:Immunohistochemistry; D009474:Neurons / Q000378:metabolism*; D010641:Phenotype; D010957:Plasmids / Q000235:genetics; D034741:RNA, Small Interfering / Q000096:biosynthesis*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection / Q000379:methods*
D005904:Glial Fibrillary Acidic Protein; D034741:RNA, Small Interfering; D049452:Green Fluorescent Proteins