实验动物雪貂阿留申病病毒核酸检测方法的建立

目的 建立阿留申病病毒PCR检测方法,以用于实验动物雪貂阿留申病病毒的检测。方法 参考Genebank中阿留申病病毒核酸VP2基因序列设计一对引物,建立PCR检测方法,进行特异性、敏感性测试,并对实验雪貂粪便样品进行筛查。结果 测序结果显示,使用设计的引物可特异性地扩增获得阿留申病病毒的VP2中531 bp基因片段;在常见实验动物DNA病毒,小鼠多瘤病毒、小鼠细小病毒、犬细小病毒、疱疹病毒、鼠痘病毒、仙台病毒、小鼠腺病毒中均未扩增出目的条带;以目的片段核酸为模板测试该方法敏感性,检测下限达到90. 6 copy/μL。应用该方法对39份雪貂粪便样品进行检测,未检测出阿留申病病毒核酸阳性。结论 本研究建立的方法具备一定的特异性和敏感性,可作为实验动物雪貂中阿留申病病毒感染病原筛查的方法。

Establishment of nucleic acid detection method for experimental animal ferret Aleutian disease virus

OBJECTIVE To establish a PCR detection method for Aleutian virus for the detection of ferrets Aleutian virus in experimental animals. Methods A pair of primers were designed with reference to the VP2 gene sequence of Aleutian disease virus in Genebank. The PCR method was established to test the specificity and sensitivity, and the experimental samples of Ferrets feces were screened. RESULTS The sequencing results showed that the designed primers can specifically amplify the 531 bp gene fragment of VP2 of Aleutian disease virus; There were no unspecific amplification when the following experimental animal DNA virus were used as templates in ADV PCR reaction: mouse Polyoma Virus, Minute virus of mice, Canine Parvovirus, Herpes Simplex virus, Ectromelia virus, Sendai virus, Mouse adenovirus; the nucleic acid plasmid with a sensitivity of 90.6 copy/μL was obtained. A total of 39 samples of ferrets were tested and no Aleutian virus nucleicacid was detected. Conclusion The current ADV PCR detection method shows applicable specificity and sensitivity, which can be used as a pathogen detection method in the screening of ferrets Aleutian disease virus in experimental animals.