摘要
目的建立一种基于多重不对称PCR-电化学芯片技术同时检测多种甲型流感病毒的新方法。方法根据常见型甲型流感病毒的基质蛋白(matrix protein,MP)、血凝素(hemagglutinin,HA)和神经氨酸酶(neuraminidase transferase,NA)基因的保守序列分别设计特异性引物和探针,建立优化多重PCR反应体系,再结合电化学芯片技术建立多重不对称PCR-电化学芯片法检测甲型流感病毒,对所建方法进行特异性和敏感性评价,同时检测200例流感样动物咽拭子样本。结果本方法检测灵敏度可达1×10~3copies/mL,200例动物样本中检出194例阳性,敏感率为97%。结论本研究建立的多重不对称PCR-电化学芯片法可同时检测常见类型甲型流感病毒,具有较高的特异性和灵敏度,且操作较简单、检测周期短,便于临床筛查、诊断和治疗。
Objective To establish a new method for simultaneous detection of multiple influenza A viruses based on multiple asymmetric PCR-electrochemical chip technology. Methods A multiplex PCR reaction system was established and optimized by designing specific primers and probes according to the conserved sequences of matrix protein（MP）,hemagglutinin（HA）and neuraminidase transferase（NA）gene encoded by influenza A viruses. The PCR system was hybridized with the chip,then,a new PCR-electrochemical chip for detecting influenza A viruses was developed. The specificity and sensitivity were evaluated. 200 samples from throat swab of animals were detected by this chip at the same time. Results The sensitivity is up to 1×10^3 copies/mL. 194 of 200 samples were detected as positive with the sensitivity of 97%.Conclusion The multiple asymmetric PCR-electrochemical chip established in this study can simultaneously detect the most common types of influenza A viruses with high specificity and sensitivity. The method of our study is simple and the detection cycle is short. It is suitable for clinical screening,diagnosis and treatment.
作者
蒋析文
黄桃生
黄志文
张险朋
殷三鸿
JIANG Xiwen;HUANG Taosheng;HUANG Zhiwen;ZHANG Xianpeng;YIN Sanhong(DAAN Gene Co., Ltd. Of Sun Yat-sen University, Guangzhou, Guangdong, China, 510665;Animal Centers for Disease Control and Prevention, Dongguan, Guangdong, China, 523000)
出处
《分子诊断与治疗杂志》
2018年第3期158-162,188,共6页
Journal of Molecular Diagnostics and Therapy
