Establishment of a fluorescence quantitative PCR assay for murine polyomavirus detection and its use in the detection in naked mole rats
10.3969.j.issn.1671-7856.2017.12.016
- VernacularTitle:小鼠多瘤病毒荧光PCR检测方法的建立及其在裸鼹鼠感染情况调查中的应用
- Author:
bo Xiao LI
1
;
Rui FU
;
jing Shu WANG
;
Li WEI
;
ming Zheng HE
;
Ji WANG
;
fei Bing YUE
Author Information
1. 中国食品药品检定研究院实验动物资源研究所
- Keywords:
Polyomavirus;
Heterocephalus glaber;
Fluorescence quantitative PCR
- From:
Chinese Journal of Comparative Medicine
2017;27(12):91-96
- CountryChina
- Language:Chinese
-
Abstract:
Objective To establish a fluorescence quantitative PCR assay for polyomavirus and to apply this technique in the investigation of its infection rate in naked mole rats. Methods To compare the nucleic acid sequence of murine polyomavirus (Genbank:NC 001515) in NCBI and design primers and probes in its conserved region. To establish a fluorescence quantitavive PCR method for polyomavirus and evaluate the sensitivity and specificity of the method. To infect nine one-day old KM strain suckling mice, and to collect samples of the heart, liver, spleen, lung, kidney, brain, thymus, cecal contents and blood at 21 days after infection. The efficacy of the method was validated by detecting the virus in organs. 62 cecal samples from naked mole rats were tested by the established assay. Results There was obvious fluorescence signal when polyomavirus was used as the template and no fluorescence signal when simian virus 40, murine K virus, MVM and H-1 were used as templates. The detection limit of the assay was 100 copies/μL. Polyomavirus DNA was detected in the heart, liver, spleen, lung, kidney and cecal contents of the mice which were inoculated with polyomavirus. The polyomavirus DNA content was highest in the lung tissue. There was no detectable polyomavirus DNA in the brain, thymus and blood of the infected mice. Sixty-two cecal contents of naked mole rats were tested for polyomavirus and the results were negative. Conclusions The fluorescence quantitative PCR assay for polyomavirus established in this study can effectively detect polyomavirus DNA in animal tissues. The results of investigation of the natural infected polyomavirus of naked mole rats provide a reference for the formulation of microbiological criteria for experimental naked mole rats.
