Analysis of Avian Influenza Virus H5 (AIV-H5) by Real-Time PCR Method (CAT#: STEM-MB-2558-LGZ)

Introduction

A real-time polymerase chain reaction (real-time PCR,or qPCR) is a laboratory technique of molecular biology based on the polymerase chain reaction(PCR). It monitors the amplification of a targeted DNA molecule during the PCR(i.e.,in real time),not at its end,as in conventional PCR.<br />Two common methods for the detection of PCR products in real-time PCR are(1)non-specific fluorescent dyes that intercalate with any double-stranded DNA and (2)sequence-specific DNA probes consisting of oligonucleotides that are labelled with a fluorescent reporter,which permits detection only after hybridization of the probe with its complementary sequence.

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Principle Applications Procedure Materials Notes Related Products

Principle

RNA was extracted by silicon matrix membrane in the centrifuge column. Under the action of reverse transcriptase, cDNA chains complementary to RNA template were synthesized using RNA as template and primers as starting point. Under the action of TaqDNA polymerase, the copy number of specific DNA fragments can be amplified by a cycle of high temperature denaturation, low temperature annealing and medium temperature extension. After 35 cycles, the amplified DNA segment was eventually magnified millions of times. The amplified DNA fragments were electrophoretic, and after staining, the amplified bands of DNA fragments were visible to the naked eye under the irradiation of ultraviolet lamp.

Applications

It is used to detect AIV-H5 in poultry tissues, secretions and excreta, and is suitable for diagnosis, detection and epidemiological investigation of AIV-H5.

Procedure

1. Sample processing and preparation of PCR reaction system.
2. Add the amplification template, cover the PCR reaction cover, mix well, centrifuge at low speed instantaneously, and transfer to the PCR instrument.
3. Set the program for PCR amplification.
4. Data analysis.

Materials

• Sample Type: serum

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