Quantitative analysis of lymphokine mRNA expression by a nonradioactive method using PCR and anion exchange chromatography.
Amplification of DNA by the polymerase chain reaction (PCR) has become an efficient tool in the study of gene expression. We describe the use of HPLC anion exchange chromatography to quantitate PCR products amplified from cDNA. The technique circumvents the use of both radioactivity and gel electrophoresis. We show that the method permits accurate quantitation of the gene product of interest and provides a clear separation of specific and non-specific products. The technique was applied to quantitate TNF-beta mRNA levels in unstimulated and stimulated mouse T cells.
Gaus
H
H
Institute of Medical Microbiology and Hygiene, Technical University of Munich, Germany.
Lipford
G B
GB
Wagner
H
H
Heeg
K
K
D000818:Animals; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange / Q000379:methods*; D004247:DNA / Q000096:biosynthesis; D015870:Gene Expression; D008233:Lymphotoxin-alpha / Q000096:biosynthesis* / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis* / Q000378:metabolism; D013601:T-Lymphocytes / Q000276:immunology; D013762:Tetrahydrofolate Dehydrogenase / Q000032:analysis; D014158:Transcription, Genetic
D008233:Lymphotoxin-alpha; D012333:RNA, Messenger; D004247:DNA; D013762:Tetrahydrofolate Dehydrogenase