T/YNANT 08-2024
Multiplex real-time fluorescence quantitative RT-PCR detection method for H5, H7, and H9 subtypes of avian influenza virus (English Version)

Standard No.

T/YNANT 08-2024

Language

Chinese, Available in English version

Release Date

2024

Published By

Group Standards of the People's Republic of China

Lastest

T/YNANT 08-2024

Scope

According to the specific sequence of the common gene of avian influenza virus, a pair of specific primers and a specific fluorescent double-labeled probe are synthesized. The 5' end and 3' end of the probe are respectively labeled with different fluorescent substances, such as the 5' end is labeled with FAM fluorescent substance, the fluorescence it emits can be received by the detection instrument, called the reporter fluorescent group (indicated by R), and the 3' end is generally labeled with TAMRA fluorescent substance, which can absorb the fluorescent signal emitted by the 5' end reporter fluorescent group in a close distance, called the quencher fluorescent group (indicated by Q). When the PCR reaction is in the annealing stage, a pair of primers and a probe bind to the target gene fragment at the same time. At this time, the fluorescent signal emitted by the R group on the probe is absorbed by the Q group, and the instrument cannot detect the fluorescent signal emitted by R; when the PCR reaction reaches the extension stage, the Taq enzyme, under the guidance of the primers, uses four nucleotides as substrates and synthesizes a new chain along the template chain according to the principle of base pairing; when the chain extension reaches the probe binding site, it is hindered by the probe and cannot continue. At this time, the Taq enzyme hydrolyzes the probe into mononucleotides to eliminate the obstruction. At the same time, the R group marked on the probe is released and received by the detector; under the action of the Taq enzyme, the extension process continues to synthesize a complete new chain, and the R and Q groups are both free in the solution, and the instrument can continue to detect the fluorescent signal emitted by R.

T/YNANT 08-2024 history

• 2024 T/YNANT 08-2024 Multiplex real-time fluorescence quantitative RT-PCR detection method for H5, H7, and H9 subtypes of avian influenza virus