科研动态

为建立一种适用于现场快速检测的猪流行性腹泻病毒（PEDV）LAMP技术，基于羟基萘酚蓝（HNB）的可视化显色特点，根据PEDV M基因编码区序列，设计合成1套引物，通过反应物浓度和反应条件优化，建立了可闭管检测的PEDV RT-LAMP检测方法。特异性和灵敏度试验结果显示，建立的RT-LAMP检测技术快速、灵敏、特异，可于1 h内检出0.2 mL 0.1 TCID50/mL的病毒RNA，与实时荧光RT-PCR检测方法灵敏度一致，与猪瘟病毒、猪伪狂犬病病毒、猪繁殖与呼吸综合征病毒、猪圆环病毒2型以及猪链球菌2型核酸不发生交叉反应。利用该方法对187份送检的粪拭子及病死猪组织样品进行应用检测，检出阳性样品9份，与荧光定量RT-PCR方法检测结果一致。试验结果表明，所建立的方法快速、特异，重复性满足要求，适用于送检样品的PEDV快速检测。

Development of HNB-based Visual RT-LAMP Method forDetection

of Porcine Epidemic Diarrhea Virus

In order to establish a RT-LAMP method forrapid detection of porcine epidemic diarrhea virus（PEDV），based on color reaction of HNB and according to M genesequence of PEDV，a set of primers were designed andsynthesized. By optimizing the reaction concentration and conditions，the RT-LAMP method was developed.Specificity and sensitivity results showed the established method was fast，sensitive and specific. It could detectPEDV-RNA extracted from 0.2 mL virus suspension（0.1 TCID50/mL），which was consistent with the real-time RT-PCR method.No cross reactions were observed with the nucleic acids of classical swinefever virus（CSFV），pseudorabies virus（PRV），porcine reproductive and respiratory syndrome virus（PRRSV），porcine circovirus type 2（PCV-2） and Streptococcus suis type 2. Using the RT-LAMPmethod，a total of 9 positive samples were detectedfrom 187 samples of fecal swabs and dead pigs submitted，which was also consistent with the resultsof real-time RT-PCR. As a conclusion，the established method was rapid，specific and repeatable，and it was suitable for  rapid detection of PEDV in submitted tissuesamples.

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