Kit for detecting five kinds of infantile diarrhea series viruses and application of kit
A technology for virus detection and infantile diarrhea, which is applied in the direction of microbial determination/inspection, DNA/RNA fragments, recombinant DNA technology, etc., can solve the problems of high reagent price, long detection time, and inability to achieve instant detection, etc., and achieve good application Effects of foreground, high sensitivity, and specificity
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Embodiment 1
[0108] Example 1 Repeatability Detection
[0109] The positive control of each virus diluted 10 times was used as the sample to be tested for repeated detection, and the detection was repeated 3 times.
[0110] 1. Nucleic acid extraction (used with Hs480 heating centrifuge)
[0111] 1) Take 50 μl of the sample and add 5 μl FastLyse L4, shake and mix;
[0112] 2) Place the mixed sample in the HS480 heating centrifuge machine, heat at 95°C for 2min, and centrifuge at 5000rpm for 2min;
[0113] 3) After centrifugation, take the supernatant as the sample RNA, numbered;
[0114] 4) The positive control, negative control and samples to be tested are processed synchronously.
[0115] 2. Preparation of PCR reagents
[0116] 1) Prepare the PCR reaction solution: pre-mix the ultra-fast PCR buffer1 (68 μl) and the ultra-fast PCR enzyme system (17 μl) (2 more than the actual number of reaction tubes, to avoid loss during sample addition, and to avoid air bubbles during mixing);
[0117...
Embodiment 2
[0134] Embodiment 2 specific detection
[0135] Each virus positive control was used as the sample to be tested for specific detection, and the detection method was the same as in Example 1.
[0136] Result: Each virus positive control sample was only positive under the corresponding probe, and the rest were negative, with good specificity. The results were as follows: Figure 1-Figure 5 shown.
Embodiment 3
[0137] Example 3 Sensitivity Detection
[0138] Each virus positive control in the kit (concentration is 1 × 10 5 copies / ml) was serially diluted to 1×10 4 copies / ml, 1×10 3 copies / ml, 1×10 2 copies / ml and 1×10 1 Copies / ml, after preparation, use it as the sample to be tested for sensitivity testing with the corresponding kit.
[0139] Concrete implementation steps are the same as embodiment 1
[0140] Results: The detection limits of NV G Ⅰ, NV G Ⅰ / G Ⅱ, HAstV, RV and EAdV were not higher than 1×10 1 copies / ml, the result is as Image 6 shown.
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