Zheng Ling-Jie, Liu Ning, Yang Kun, Wang Ai-Feng, Tan Zhi-Rong, Li Xiang
1 Fuwai Central China Cardiovascular Hospital, Zhengzhou, China.
2 Coyote Bioscience Co., Ltd., Haidian District, Beijing, China.
J Int Med Res. 2018 Dec;46(12):4965-4973. doi: 10.1177/0300060518787718. Epub 2018 Oct 25.
To directly achieve cytochrome P450 2C19 gene ( CYP2C19) classification using one-step real-time fluorescent PCR detection and to verify the capabilities of this method with nucleic acid extracted from whole blood samples.
A human CYP2C19 genotyping kit based on one-step real-time fluorescent PCR detection was used to analyze whole blood or genomic DNA samples. This method was compared with pyrosequencing and another quantitative (q)PCR kit for its accuracy, repeatability, detection range analysis, sensitivity, specificity, and anti-interference analysis.
The one-step real-time PCR method achieved a 100% accuracy rate compared with pyrosequencing and the other qPCR kit. When detecting different concentrations of known genes, concentrations of each sample ranging from 0.2 to 125 ng/µL could be correctly detected. The genotypes of samples treated with anticoagulants, including EDTA and sodium citrate, and chyle blood samples could be correctly detected.
The one-step detection method demonstrated high accuracy and a wide detection range. It also had high levels of repeatability, sensitivity, and specificity for the assessment of genomic DNA test samples.
采用一步法实时荧光定量聚合酶链反应（PCR）检测直接实现细胞色素P450 2C19基因（CYP2C19）分型，并利用全血样本提取的核酸验证该方法的性能。
使用基于一步法实时荧光定量PCR检测的人CYP2C19基因分型试剂盒分析全血或基因组DNA样本。将该方法与焦磷酸测序法和另一种定量PCR试剂盒在准确性、重复性、检测范围分析、灵敏度、特异性及抗干扰分析等方面进行比较。
与焦磷酸测序法和另一种定量PCR试剂盒相比，一步法实时荧光定量PCR检测法的准确率达100%。检测不同浓度的已知基因时，可正确检测出浓度范围在0.2至125 ng/µL的各样本。使用包括乙二胺四乙酸（EDTA）和柠檬酸钠在内的抗凝剂处理过的样本以及乳糜血样本的基因分型均可被正确检测。
一步法检测具有较高的准确性和较宽的检测范围。在评估基因组DNA检测样本时，该方法还具有高度的重复性、灵敏度和特异性。
