Detection of measles virus infection and proliferation by real-time quantitative PCR
10.19428/j.cnki.sjpm.2021.19441
- VernacularTitle:实时定量PCR技术检测麻疹病毒的感染与增殖
- Author:
Ran XU
1
;
Lin-ya ZHANG
;
Wen-zhen YANG
;
Wei ZHU
Author Information
1. Research Laboratory, Shanghai Institute of Biological Products Co., Ltd., Shanghai 200051, China
- Publication Type:Research Article
- Keywords:
real-time quantitative PCR;
SYBR-Green;
measles virus N gene
- From:
Shanghai Journal of Preventive Medicine
2021;33(2):132-
- CountryChina
- Language:Chinese
-
Abstract:
Objective A SYBR-Green real-time quantitative PCR(RT-qPCR)method was set up to detect the infection and proliferation of measles virus, which could be useful in virus titer determination. Methods In this project, we used a 405 bp fragment of the N gene of measles virus as a target sequence and constructed a plasmid to establish the standard curve in absolute quantitative experiment. We then used this method to obtain the proliferation curve of measles virus and to detect the virus proliferation at different MOI. Results There was a linear relationship between the virus copy number and the titer of the measles virus reference at the range of 6 to 2 lgCCID50/mL, with a correlation coefficient (r) of 0.991(P < 0.01). Based on the analysis of virus proliferation curve, measles virus mainly proliferated intracellularly within 48 h after its entering the cell. There was no detected increase in viral RNA level in the first 24 h, suggesting the virus was in a silent period in the cell. After 24 h, the virus expanded in large numbers and entered the exponential growth phase. The intracellular viral RNA level reached the plateau phase after its peak at 96 h. The virus secreted to the outside of the cell entered the exponential growth phase starting from 48 h, peaked at 144 h, then followed by plateau phase. Conclusion A SYBR-Green RT-qPCR method is established and used to monitor virus proliferation. Our result is helpful in understanding of the proliferation and secretion of measles virus in cells and provides experimental basis for detection of live attenuated virus titers.
