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Genotyping of K-ras codons 12 and 13 mutations in colorectal cancer by capillary electrophoresis
- Source :
-
Journal of Chromatography A . Jun2009, Vol. 1216 Issue 26, p5147-5154. 8p. - Publication Year :
- 2009
-
Abstract
- Abstract: Point mutations of the K-ras gene located in codons 12 and 13 cause poor responses to the anti-epidermal growth factor receptor (anti-EGFR) therapy of colorectal cancer (CRC) patients. Besides, mutations of K-ras gene have also been proven to play an important role in human tumor progression. We established a simple and effective capillary electrophoresis (CE) method for simultaneous point mutation detection in codons 12 and 13 of K-ras gene. We combined one universal fluorescence-based nonhuman-sequence primer and two fragment-oriented primers in one tube, and performed this two-in-one polymerase chain reaction (PCR). PCR fragments included wild type and seven point mutations at codons 12 and 13 of K-ras gene. The amplicons were analyzed by single-strand conformation polymorphism (SSCP)-CE method. The CE analysis was performed by using a 1× Tris–borate–EDTA (TBE) buffer containing 1.5% (w/v) hydroxyethylcellulose (HEC) (MW 250 000) under reverse polarity with 15°C and 30°C. Ninety colorectal cancer patients were blindly genotyped using this developed method. The results showed good agreement with those of DNA sequencing method. The SSCP-CE was feasible for mutation screening of K-ras gene in populations. [Copyright &y& Elsevier]
Details
- Language :
- English
- ISSN :
- 00219673
- Volume :
- 1216
- Issue :
- 26
- Database :
- Academic Search Index
- Journal :
- Journal of Chromatography A
- Publication Type :
- Academic Journal
- Accession number :
- 41242296
- Full Text :
- https://doi.org/10.1016/j.chroma.2009.04.083