Triple fluorescence quantitative reverse transcription-polymerase chain reaction (RT-PCR) detection kit and use thereof
A detection kit and real-time fluorescence quantitative technology, applied in the field of life, can solve the problems of wasting clinical specimens and reagents, time-consuming and labor-intensive expenses, and achieve the effect of reducing mortality and sequelae
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Embodiment 1
[0050] see figure 1 , a triple real-time fluorescence quantitative RT-PCR detection kit, including: quantitative RT-PCR reaction solution, Zyzavirus standard, Astrovirus standard, Adenovirus standard, positive control, negative control, box body 7. There are container holes for placing the quantitative RT-PCR reaction solution tube 1, the standard sample tube 2 of Zaravirus, the standard tube 3 of astrovirus, the 4 standard sample of adenovirus, the positive control tube 5, and the negative control tube 6. , wherein the quantitative RT-PCR reaction solution is packed in a single tube, arranged in a matrix, and the standard products are gene standards of Sarsovirus, Astrovirus and adenovirus, wherein the fluorescent quantitative RT-PCR reaction solution contains RT-PCR reaction buffer (containing Magnesium chloride and deoxyribonucleotide triphosphate mixture, etc.) 3 tubes (labeled as ), three virus universal primers (upstream and downstream primers in the same tube) are 3 ...
Embodiment 2
[0052] 1 Materials and methods
[0053] Virus strains and clinical specimens:
[0054] The positive nucleic acids of enteric adenovirus 40 / 41, astrovirus and saruvirus (all identified by gene sequencing) were from the First Affiliated Hospital of Zhejiang University School of Medicine. Clinical samples were obtained from patients' stool samples, which were collected and transported to the laboratory on ice.
[0055] 1.2 Primers and probes
[0056] Downloaded the gene sequences of Sarsovirus, Astrovirus, and Adenovirus from all over the world from the NCBI Gene Bank in the United States. The homology comparison was carried out, and specific primers and Taqman probes were designed in the conserved gene region corresponding to the virus genome. The sequence is as follows:
[0057] Upstream primer binding such as virus-FP: 5'- CAACTATGACCAGGCTCTCGC-3'
[0058] Downstream primer binding such as virus-RP: 5'- GCCCTCCATYTCRAACACTA-3'
[0059] Specific probes such as virus-P: 5'-...
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