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多重PCR-MALDI-TOF MS技术用于鉴别奥密克戎EG.5和BA.2.86变异株

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鉴于奥密克戎EG.5 和BA.2.86 变异株的强传播能力,本研究将基质辅助激光解吸电离飞行时间质谱与多重PCR技术联用,针对其刺突(S)蛋白上的关键突变位点进行扩增和延伸引物设计,同时引入用于鉴别的D614G位点和用于SARS-CoV-2鉴定的ORF1ab及N基因检测位点,可一次性检测10个特征位点.结果表明,该方法可在同一反应体系中同时检测2种重点关注变异株EG.5和BA.2.86,检测结果准确,灵敏度低至20 copies/反应.该方法适合海关及其他疫情监测部门对重点关注变异株进行大量样本的动态筛查工作,是荧光定量PCR和高通量测序方法很好的技术补充,可为病毒溯源和疫情监测提供技术支撑.
The Application of Multiplex PCR-MALDI-TOF MS Technique for Identification of Omicron EG.5 and BA.2.86 Variants
In light of the highly transmissible nature of the Omicron EG.5 and BA.2.86 variants,this study integrates Matrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry(MALDI-TOF MS)with multiplex PCR technology.Primers were designed to amplify and extend key mutation sites on the spike(S)protein of these variants.Additionally,the D614G mutation site was included for identification,along with the ORF1ab and N gene detection sites for the identification of SARS-CoV-2,enabling the simultaneous detection of 10 characteristic sites.The results demonstrate that this method can detect both the EG.5 and BA.2.86 variants in the same reaction system with high accuracy and a sensitivity as low as 20 copies per reaction.This method is well-suited for dynamic screening of a large number of samples by customs and other epidemic monitoring departments,focusing on variants of concern.It serves as a valuable technical complement to quantitative fluorescent PCR and high-throughput sequencing methods,providing technical support for virus tracing and epidemic surveillance.

Omicron variantsmultiplex PCRMatrix-Assisted Laser Desorption Ionization Time-of-Flight Mass Spectrometry(MALDI-TOF MS)typing screening

张晓龙、魏海燕、郭悠然、袁芳、祁科元、郭惠民、宋悦谦

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中国海关科学技术研究中心 北京 100026

浙江迪谱诊断技术有限公司 杭州 311100

奥密克戎变异株 多重PCR 基质辅助激光解吸电离飞行时间质谱 分型筛查

海关总署科研项目

2022HK149

2024

中国口岸科学技术
中国质检报刊社

中国口岸科学技术

影响因子:0.051
ISSN:1002-4689
年,卷(期):2024.6(8)
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