Abstract
This study aimed to develop a simple and rapid method to detect KRAS gene mutations for conventional clinical applications under laboratory conditions. The genotype of mutation sites was determined based on the occurrence of target bands in the corresponding lanes of the reaction tubes through polymerization-conjunction of the probes, probe purification and amplification, and agarose gel electrophoresis. Circulating DNA samples were obtained from the plasma of 72 patients with lung cancer, which were identified based on six mutation sites (G12S, G12R, G12C, G12D, G12A, and G12V) of codon 12 of the KRAS gene. The detection results were compared with direct sequencing data. The proposed detection method is characterized by simple operation, high specificity, and high sensitivity (2%). This method can detect the mutations of three samples at G12S, G12R, and G12A. In the direct sequencing spectra of these samples, the genotype could not be determined due to the lack of evident sequencing peaks that correspond to the basic group of mutations. In conclusion, a simple and rapid method was established based on probe polymerization-conjunction-agarose gel electrophoresis for detecting KRAS gene mutations. This method can be applied to the conventional mutation detection of inhomogeneous samples.
Highlights
The KRAS gene is a common oncogene in humans, and encodes the 21-kDa RAS protein
The present study established a simple and fast method for KRAS gene mutation detection based on the method for detecting epidermal growth factor receptor (EGFR) gene mutations (Tang et al, 2014)
The probe length is less than 50 bp, and it can detect mutation alleles using ordinary PCR instruments and agarose gel electrophoresis (AGE)
Summary
The KRAS gene is a common oncogene in humans, and encodes the 21-kDa RAS protein. The RAS protein exhibits intrinsic guanosine triphosphate (GTP) enzymatic activity, and is involved in cell proliferation, differentiation, and apoptosis by switching the mutual transformation regulatory signal system between guanosine diphosphate (GDP) and GTP. The KRAS gene has a high mutation rate in patients with colorectal cancer and lung cancer, and confers the resistance to epidermal growth factor receptor (EGFR), tyrosine kinase inhibitors (TKIs), and EGFR monoclonal antibody agents (Chen et al, 2013; Li et al, 2014; Leiser et al, 2015; Hsu et al, 2016). The mutation rate of the KRAS gene can reach as high as 15–30% in Caucasian patients with colorectal cancer, and only 4–10% in Asian patients with lung cancer (Mascaux et al, 2005; Lu et al, 2013; Kinugasa et al, 2015; Omidifar et al, 2015; Ohba et al, 2016). The detection of KRAS gene mutations is important for the development of individualized treatments for patients with tumors, as well as to improve the effects of targeted clinical treatments and reduce treatment expenses
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
