M-gene based fluorescent RT-PCR (Reverse Transcription Polymerase Chain Reaction) detection method of Nipah virus
A RT-PCR, Nipah virus technology, applied in the field of inspection and quarantine, can solve the problems of no effective treatment and prevention, Nipah disease threat, etc., to achieve the effects of simple operation, enhanced specificity, and high specificity
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[0043] 1. Sample processing and RNA extraction
[0044] Take 100mg of pig product tissue and grind it for RNA extraction.
[0045] Take 200 μL of the pseudovirus and add it to an EP tube containing 750 μL Trizol reagent without RNase, shake it well for 15 seconds, and let it stand at room temperature for 5 minutes. Add 200 μL of RNase-free chloroform, shake for 15-30 seconds, let stand at room temperature for 2-3 minutes, centrifuge at 12,000 r / min at 4°C for 15 minutes; carefully absorb the upper aqueous phase and transfer it to a new EP tube; add unused RNase-free iso Propanol 500μL, mix well, let stand at room temperature for 10min; centrifuge at 4℃, 12000r / min for 10min; discard the supernatant, add 1mL of 75% ethanol prepared with DEPC water, shake, fully wash the precipitate, 4℃, 12000r / min Centrifuge at 1 / min for 5 min; discard the supernatant, dry in vacuum, add 5-10 μL RNase-free triple distilled water to dissolve the precipitate, and set aside. Take the ground 100m...
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