Multiplex reverse transcription-polymerase chain reaction combined with on-chip electrophoresis as a rapid screening tool for candidate gene sets.
Combining multiplex reverse transcription-polymerase chain reaction (mRT-PCR) with microfluidic amplicon analysis, we developed an assay for the rapid and reliable semiquantitative expression screening of 11 candidate genes for drug resistance in human malignant melanoma. The functionality of this approach was demonstrated by low interexperimental variations of amplicon quantities after endpoint analysis. When applied to RNA samples derived from drug-sensitive and -resistant melanoma cell lines, mRT-PCR delivered results qualitatively concordant with data obtained from Northern blot and array analyses. The screening of additional melanoma cell lines resulted in distinct expression patterns for ten candidate genes. Our approach reveals a rapid and easy-to-handle alternative for candidate gene set evaluation from limited amounts of RNA.
Wittig
Rainer
R
Molecular Genome Analysis, Deutsches Krebsforschungs-zentrum (DKFZ), Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany. r.wittig@dkfz.de
Salowsky
Rüdiger
R
Blaich
Stephanie
S
Lyer
Stefan
S
Maa
Juehn S
JS
Müller
Odilo
O
Mollenhauer
Jan
J
Poustka
Annemarie
A
D045744:Cell Line, Tumor; D019008:Drug Resistance, Neoplasm / Q000235:genetics*; D046209:Electrophoresis, Microchip; D005820:Genetic Testing / Q000379:methods*; D006801:Humans; D008545:Melanoma / Q000175:diagnosis / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*