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- Odin, Elisabeth, 1955, et al.
(author)
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Rapid method for relative gene expression determination in human tissues using automated capillary gel electrophoresis and multicolor detection
- 1999
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In: J Chromatogr B Biomed Sci Appl. - 1387-2273. ; 734:1, s. 47-53
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Journal article (peer-reviewed)abstract
- The aim of this study was to evaluate a direct and automated post-polymerase chain reaction (PCR) detection system to simultaneously determine the relative gene expression levels of nine cancer-related human genes. Total RNA was prepared from flash-frozen biopsies derived from human colorectal tumors or normal mucosa and reverse-transcribed to cDNA which was PCR-amplified using primer pairs corresponding to the studied genes. In each reaction, the forward primer was labeled with a fluorescent dye. The PCR products were pooled and an internal size standard with a uniquely colored fluorescent dye was added. The samples were then subjected to automated capillary gel electrophoresis. Fragment analysis software was used to calculate the relative gene expression using beta-actin as the reference gene. We found that automated capillary gel electrophoresis with multicolor detection is a rapid, accurate and highly reproducible method for separation and quantification of PCR-amplified cDNA.
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