| 宫艳艳,马 娟,刘 英,陶志华,张 钧,崔 康.双通道TaqMan实时荧光PCR应用于别嘌呤醇不良反应相关基因HLA-B*58:01的检测与验证[J].现代生物医学进展英文版,2022,(21):4163-4167. |
| 双通道TaqMan实时荧光PCR应用于别嘌呤醇不良反应相关基因HLA-B*58:01的检测与验证 |
| Detection and Validation of Allopurinol Adverse Reaction Related Gene HLA-B * 58:01 by Dual Channel TaqMan Real-Time Fluorescent PCR |
| Received:May 20, 2022 Revised:June 15, 2022 |
| DOI：10.13241/j.cnki.pmb.2022.21.030 |
| 中文关键词: 别嘌呤醇 HLA-B*58:01 基因检测 Taqman 探针 实时荧光 PCR |
| 英文关键词: Allopurinol HLA-B*58:01 Gene detection Taqman probe Real-time fluorescence PCR |
| 基金项目:陕西省重点研发项目（2019SF-188） |
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| 中文摘要: |
| 摘要 目的：建立一种双通道TaqMan实时荧光PCR方法，应用于别嘌呤醇不良反应相关基因HLA-B*58:01的快速基因检测，并验证该方法的灵敏度、特异性、重复性及可靠性等性能指标。方法：针对HLA-B*58:01 等位基因序列设计引物和Taqman 探针，建立一种双通道TaqMan实时荧光PCR检测方法，收集陕西省人民医院、浙江大学医学院附属第二医院、浙江大学医学院附属邵逸夫医院三家医院收治的1158例临床诊断为痛风或血清尿酸高患者的外周血标本，采用双盲对照实验，分别采用双通道TaqMan实时荧光PCR法与已获审批的商业试剂盒共同检测所有标本，对比两种方法的检测结果进而评价该TaqMan实时荧光PCR方法的临床性能。结果：双通道TaqMan实时荧光PCR法特异性高，稳定可靠，可检测低至1 ng/?滋L的阳性样本；采用双通道TaqMan实时荧光PCR法与已获审批的商业试剂盒在1158份患者标本中检出的HLA-B*58:01阳性及阴性标本均相同，阳性标本147例，阴性标本1011例，该组患者HLA-B*58:01基因携带率为12.69%，两种方法符合率为100%（1158/1158），两组实验结果数据采用SPSS分析P=1.000，差异无统计学意义。所有受检者HLA-B* 58:01携带率为12.69 % （147 /1158）。结论：双通道TaqMan实时荧光PCR可以作为一种快速、特异、灵敏的HLA-B*58:01的基因检测方法，用于别嘌呤醇用药前的相关不良反应风险评估。 |
| 英文摘要: |
| ABSTRACT Objective: A dual channel TaqMan real-time fluorescent PCR method was developed for the rapid detection of HLA-B *58:01 genes associated with the adverse reaction of allopurinol. The sensitivity, specificity, repeatability and reliability of the method were verified. Methods: Primers and Taqman probes were designed for HLA-B*58:01 allele sequences to establish a dual channel TaqMan real-time fluorescent PCR detection method. A double-blind controlled experiment was conducted to collect peripheral blood samples from 1158 patients clinically diagnosed with gout or high serum uric acid who were admitted to Shaanxi Provincial People's Hospital, the Second Affiliated Hospital of Zhejiang University School of Medicine, and Shaw Hospital Affiliated to Medical College of Zhejiang University. Dual channel TaqMan real-time fluorescent PCR was used to detect all specimens together with the approved commercial kit, and the results of the two methods were compared to evaluate the clinical performance of the TaqMan real-time fluorescence PCR method. Results: Dual channel TaqMan real-time fluorescent PCR method has high specificity, stability and reliability, and can detect positive samples as low as 1 ng/μL. The positive and negative HLA-B*58:01 samples were the same in 1158 patient samples by dual channel TaqMan real-time fluorescent PCR and approved commercial kit, with 147 positive samples, and 1011 negative samples. The carrying rate of HLA-B*58:01 gene in this group was 12.69%. The coincidence rate of the two methods was 100% (1158/1158). SPSS was used to analyze the experimental data of the two groups, P=1.000, and the difference was not statistically significant. The prevalence of HLA-B* 58:01 in all subjects was 12.69 % (147/1158). Conclusion: The dual channel TaqMan real-time fluorescent PCR can be used as a rapid, specific and sensitive HLA-B*58:01 gene detection method, it is used for the risk assessment of related adverse reactions before allopurinol administration. |
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