中国农业科学院农业信息研究所 中国农学会
Agricultural Information Institute of CAAS China Association of Agricultural Science Societies
En
Ch
中国农业科学院农业信息研究所 中国农学会 主办

AI检索

订阅

返回顶部
当前位置: 首页 > 文章 > 施马伦贝格病焦磷酸测序检测方法的建立 中国动物检疫 2014 (10) 57-61+65
Position: Home > Articles > Development of PCR-Pyrosequencing Assay for Detection of Schmallenberg Disease China Animal Health Inspection 2014 (10) 57-61+65

施马伦贝格病焦磷酸测序检测方法的建立

作  者：
王彩霞;林祥梅;张永宁;吕继洲;冯春燕;袁向芬;邓俊花;吴绍强
单  位：
中国检验检疫科学研究院动物检疫研究所
关键词：
施马伦贝格病毒;聚合酶链反应;焦磷酸测序;鉴定;检测
摘  要：
为适应口岸施马伦贝格病快速、准确、高通量检测需求,建立一种基于RT-PCR及焦磷酸测序技术平台的施马伦贝格病检测方法。本研究针对施马伦贝格病毒基因组RNA的S、M和L三个基因节段保守区域利用焦磷酸测序软件PyroMark Q96ID分别设计RT-PCR扩增引物及焦磷酸测序引物,以人工合成的SBV重组质粒为模板分别建立了SBV S、M、L基因的PCR-焦磷酸测序检测方法,比较三基因的PCR扩增结果以及测序分析结果,结合特异性检测结果,最终确定以M基因作为靶基因建立的焦磷酸测序检测方法效果最好,并以德国FLI提供的SBV(BH80株)RNA对所建立的方法进行验证,通过对测得序列的比对分析可确定为施马伦贝格病毒,这表明本研究所建立的方法灵敏度高、稳定性好、特异性强,可以用于施马伦贝格病毒基因序列水平上的精准检测鉴定。
译  名：
Development of PCR-Pyrosequencing Assay for Detection of Schmallenberg Disease
作  者：
Wang Caixia;Lin Xiangmei;Zhang Yongning;Lv Jizhou;Feng Chunyan;Yuan Xiangfen;Deng Junhua;Wu Shaoqiang;The Institute of Animal Quarantine,Chinese Academy of Inspection and Quarantine;
关键词：
Schmallenberg disease;;polymerase chain reaction;;pyrosequencing;;identifi cation;;detection
摘  要：
To meet the rapid,accurate and high throughput detection of Schmallenberg disease,a pyrosequencing assay was developed in combination with PCR amplifi cation of the target sequences. Pyrosequencing specifi c primers of SBV segments of S,M and L were separately designed targeting the conserved region of the sequence using primer designer software PyroMark Q96 ID. PCR-pyrosequencing assay of SBV S,M,L were established with the recombinant plasmid of SBV S,M,L as template,and the sequences were determined by PyroMarkTM ID System. SBV RNA presented by FLI was used as positive material to validate the developed PCR-pyrosequencing assay. Comparing the electrophoresis result of PCR amplicons,the result of pyrosequencing assay and the result of specifi city detection,the pyrosequencing assay targeting M gene was the best one among the assays targeting S,M and L genes. The developed SBV-M pyrosequencing assay was further verifi ed by using the SBV positive RNA material as template by means of Blast analysis of obtained sequence online. Results showed that the developed pyrosequencing assay could identify SBV. The method was of high sensitivity,good stability and high specifi city,and could be used as an accurate detection method for Schmallenberg disease.

相似文章

导出引用
计量
文章访问数: 11
HTML全文浏览量: 0
PDF下载量: 0

所属期刊

推荐期刊

友情链接Friendly link
农业信息研究所Agricultural information institute 农业智能知识服务平台Agricultural intelligent knowledge service platform 农业学术服务平台Agricultural Knowledge Service System 中国知网CNKI 维普网VIP 万方WANFANG DATA
联系我们Contact us

电话Tell：010-82109652-808

邮箱E-mail：agrijournal@caas.cn

服务Service
关于我们About us 意见建议Opinions and suggest
版权说明Copyright
单位：中国农业科学院农业信息研究所 中国农学会 © 2017中国农业科学院农业信息研究所 ICP备案号：京ICP备09089781号-46