An Extraction-Free Method for Rapid Detection of CYP2C19 * 2/3/17
Polymorphisms in One Tube using Melting Curve Analysis
Abstract
Drug-metabolizing enzymes play an important role in the metabolism of
drugs in vivo. Their activity is an important factor affecting the rate
of drug metabolism, which directly determines the intensity and
persistence of drug action. Patients taking medication can be divided
into different metabolic types through detection of CYP2C19
drug-metabolizing enzyme gene polymorphisms, which can then be used for
medication guidance for clopidogrel. Here, we describe a detection
method based on real-time polymerase chain reaction. This method uses
multicolor melting curve analysis to accurately identify different
mutation sites and genotypes of CYP2C19 * 2, CYP2C19 * 3, and CYP2C19 *
17. The detection limit of plasmid samples was 1 copies/µl; that of
genomic samples was 0.1 ng/µl. The system can detect nine types of
CYP2C19 * 2/3/17 at three sites in one tube, quickly achieving detection
within 1 h. Combined with the sample release agent, sample extraction
was completed in 5 s, achieving rapid diagnosis without extraction for
timely diagnosis and treatment. Furthermore, the system is not limited
to blood samples and can also be applied to oropharyngeal and saliva
samples, increasing sampling diversity and convenience. When using
clinical blood samples (n=93), the detection system we established was
able to quickly and accurately identify different genotypes, and the
accuracy and effectiveness of the detection were confirmed by Sanger
sequencing.