[Rapid detection of the N1 and N2 subtype of influenza virus by duplex fluorescence quantitative RT-PCR].
OBJECTIVE
To develop a method to rapidly detect the N1 and N2 subtype of influenza virus by fluorescence real-time quantitative PCR.


METHODS
According to the conservative sequences of NA gene of the N1 and N2 subtype of influenza virus, two pairs of primer and Taq-man probe were designed, respectively. Using one step RT-PCR kit to set up a duplex RT-PCR system for detecting the N1 and N2 subtype of influenza virus; the standard quantitative curve of the assay was established by using 10-fold serial dilution of in-vitro transcribed RNA, and the sensitivity and reproducibility was determined. The specificity and test for influenza virus and respirovirus were also determined by using this duplex Real-time RT-PCR system.


RESULTS
The sensitivity of detecting the N1 and N2 subtype of influenza virus was 10 copies/microl, and the regression coefficient of the quantitative curve was higher than 99%. The amplification efficiency and specificity of this assay was 102.21% and 101.78% respectively. The system was capable of detecting human influenza viruses with high specificity.


CONCLUSIONS
The detection system based on RT-PCR could be utilized to rapidly and sensitively detect the N1 and N2 subtype of influenza virus.
Liang
L
Huang
Hongyu
H
Zhang
Ting
T
Wang
Shisong
S
Fang
Xin
X
Wang
Jianxiong
J
Li
Xiaowen
X
Cheng
Xing
X
Lv
Chunli
C
Wu
Renli
R
Zhang
Jinquan
J
Cheng
D017931:DNA Primers; D005453:Fluorescence; D006801:Humans; D009980:Influenza A virus / Q000302:isolation & purification*; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017931:DNA Primers