Rt-Qpcr Detection Kit

Description This kit utilizes fluorescence quantitative probe-based PCR and guarantees a high specificity to ensure accurate one-step identification of ORF1ab and N genes of 2019-nCoV. The kit offers a highly sensitive test with a limit of detection as low as 500 copies/ml. The results are available within 1.5 hour. Kit Contents (48 Tests /Kit) Dual Enzyme solution A 50 µl Dual Ncvo-O / N reaction solution A 1 ml Dual Ncvo-O / N positive control A 200 µl Dual Negative control A 200µl Applicable equipment ABI series, Bio-Rad, Roche series and other fluorescent real-time PCR instruments. Specimens requirements Specimen type: nasopharyngeal swab from suspected infection; virus cell culture fluid, etc. Basic Protocol 1. Sample preparation. Take the test sample and extract the RNA nucleic acid according to the instructions of the nucleic acid extraction kit. The nucleic acid extraction product should be stored at -20 C. 2. Reaction mixture preparation According to the total number of test samples, the number of PCR reaction tubes needed is. N = number of samples + 1 negative control + 1 positive control. The following protocol is recommended for a 20 µl reaction volume. If the volume of reaction changes, please adjust proportionally. Components Volume Dual Enzyme solution A 1 µl Dual Ncvo-O / N reaction solution A 19µl Final Volume: 20 µl 3. Sample adding. Add 5 µl each of the extracted RNA, positive and of negative control to corresponding reaction tubes. After assembling all the components, cover the tube caps and gently mix the contents of the tube, mix well, and centrifuge briefly. 4. Perform quantitative PCR Place the reaction tube inside a real-time PCR instrument. Set the channel and sample information, reaction system volume 25 µl. Select the following channels: FAM channel for nCOV-ORF1ab, VIC channel for nCOV-N. Perform quantitative PCR using recommended cycling parameters settings: Step Temperature Time Number of cycles Fluorescence Detector 1 50C 15 min 1 Off 2 95C 3 min 1 Off 3 95C 10sec 45 Off 60C 30 sec On Result analysis: 1) Set the baseline: Generally, it is set to 6-15cycle for ABI 7500, 7700 and other instruments, 3-15cycle for PE5700, and 6-12cycle for MJ Research Option2. Under special circumstances, the baseline can be adjusted accordantly. 2) Set the threshold: The threshold line just exceeds the highest point of the negative control amplification curve (random noise line).

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RT PCR-Detection Kit