鼠痘病毒FQ-PCR检测方法的建立及初步应用  

Development of an FQ-PCR method to detect and quantitate ectromelia virus and primary applications

作　　者：王莎莎[1] 付瑞[1] 王吉[1] 岳秉飞[1] WANG Shasha;FU Rui;WANG Ji;YUE Bingfei(National Institute for Food and Drug Control,Beijing 102629,China)

机构地区：[1]中国食品药品检定研究院,北京102629

出　　处：《中国比较医学杂志》2021年第1期101-105,共5页Chinese Journal of Comparative Medicine

基　　金：国家科技支撑计划(2015BAI09B02)。

摘　　要：目的建立鼠痘病毒(ECTV)的Taq Man探针法荧光定量PCR检测方法,并对实验室保存的裸鼹鼠组织样本、小鼠组织样本以及黄鼠组织样本进行检测。方法根据Gen Bank中鼠痘病毒的crm D基因序列设计引物探针,建立ECTV的FQ-PCR检测方法,对保存的样本进行筛查,并对部分阳性样本进行测序鉴定。结果建立的ECTV FQ-PCR检测方法灵敏度高、特异性强,最低能检测到ECTV核酸浓度为10 copies/μL的样本,方法的标准曲线相关参数符合标准。使用该方法检测实验室63份裸鼹鼠脾组织样本和22份小鼠脾组织样本,结果全为阴性,检测4只黄鼠组织样本,结果阳性率为50%。阳性样本进行PCR检测后测序比对为ECTV核酸序列。结论经验证,建立的ECTV FQ-PCR检测方法能够灵敏特异的对样本中的ECTV进行筛查,实验室小鼠的日常监测不应忽视ECTV污染的潜在风险。Objective To identify the prevalence of ECTV in mice.Methods A sensitive and specific FQ-PCR assay for ECTV was developed and used to detect and quantitate ECTV in mouse samples by analyzing the sequence of the crm D gene in ECTV.Results This assay detected a minimum of 10 copies of standard DNA.The assay was applied to 63 naked mole rat spleen samples and 22 mouse spleen samples raised in the laboratory,which provided negative result.In tissue samples from four ground squirrels raised in our laboratory,the positive rate was 50%.Some positive samples amplified with primers for the crm D gene had 99%similarities to ECTV as determined by sequencing the PCR products.Conclusions Our result suggest that routine surveillance of ECTV in laboratory mice cannot be ignored.

关 键 词：鼠痘病毒 TaqMan探针法荧光定量PCR方法 裸鼹鼠 黄鼠 

分 类 号：R-33[医药卫生]