TaqMan Real-time RT-PCR Assay for Detecting and Differentiating Japanese Encephalitis Virus / 生物医学与环境科学(英文)
Biomedical and Environmental Sciences
; (12): 208-214, 2018.
Article
in En
| WPRIM
| ID: wpr-690669
Responsible library:
WPRO
ABSTRACT
<p><b>OBJECTIVE</b>To detect Japanese encephalitis virus (JEV) rapidly and distinguish its genotypes, a TaqMan-based reverse transcriptase quantitative polymerase chain reaction (RT-PCR) detection system was developed.</p><p><b>METHODS</b>By aligning the full-length sequences of JEV (G1-G5), six sets of highly specific TaqMan real-time RT-PCR primers and probes were designed based on the highly conserved NS1, NS2, and M genes of JEV, which included one set for non-specific JEV detection and five sets for the detection of specific JEV genotypes. Twenty batches of mosquito samples were used to evaluate our quantitative PCR assay.</p><p><b>RESULTS</b>With the specific assay, no other flavivirus were detected. The lower limits of detection of the system were 1 pfu/mL for JEV titers and 100 RNA copies/µL. The coefficients of variation of this real-time RT-PCR were all < 2.8%. The amplification efficiency of this method was between 90% and 103%.</p><p><b>CONCLUSION</b>A TaqMan real-time RT-PCR detection system was successfully established to detect and differentiate all five JEV genotypes.</p>
Key words
Full text:
1
Index:
WPRIM
Main subject:
Virology
/
Polymerase Chain Reaction
/
Reproducibility of Results
/
Sensitivity and Specificity
/
Encephalitis Virus, Japanese
/
Genetics
/
Methods
/
Culicidae
Type of study:
Diagnostic_studies
Limits:
Animals
Language:
En
Journal:
Biomedical and Environmental Sciences
Year:
2018
Type:
Article