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Universal nucleic acid detection kit and method for human adenoviruses

A detection kit, human adenovirus technology, applied in the direction of microorganism-based methods, biochemical equipment and methods, microorganism measurement/testing, etc., can solve the problems of insensitivity of clinical specimens, susceptibility to bacterial and fungal contamination, etc. Achieve the effect of mature detection method, good specificity and reliable experimental basis

Pending Publication Date: 2020-08-18
嘉兴实践医学科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although cell culture remains the gold standard for HAdV, it is insensitive to clinical specimens, slow and susceptible to bacterial and fungal contamination

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] A human adenovirus universal nucleic acid detection kit, which includes an extraction solution for extracting human adenovirus DNA, a PCR reaction solution, and the reaction solution includes 50 μL of 13 μM Taq DNA polymerase, 100 μL of 150 μM dNTP, 200 μL of SyBr dye, and 15 μM of uracil glycosyl Lyase 50 μL, human adenovirus positive control substance, negative control substance;

[0023] The primer sequences used in the DNA amplification reaction in the PCR reaction solution are as follows:

[0024] P1: 5'-ACATCGCCGGACAGGATGCTTCGGAGTACC-3';

[0025] P2: 3'-CACCCGCTGTTGTCTCACGACCTATACCGG-5'.

[0026] The PCR reaction system is 20 μL, including 15 μL of the PCR reaction solution, 2 μL of the primers and 3 μL of sample DNA;

[0027] The PCR reaction cycle parameters are:

[0028] 94°C, 2min; enter the cycle stage: denaturation at 94°C for 15s, annealing at 50°C for 1min, extension at 72°C for 20s, same as 35 cycles; stop at 10°C.

Embodiment 2

[0030] A human adenovirus universal nucleic acid detection kit, which includes an extraction solution for extracting human adenovirus DNA, a PCR reaction solution, and the reaction solution includes 50 μL of 13 μM Taq DNA polymerase, 100 μL of 150 μM dNTP, 200 μL of SyBr dye, and 15 μM of uracil glycosyl Lyase 50 μL, human adenovirus positive control substance, negative control substance;

[0031] The primer sequences used in the DNA amplification reaction in the PCR reaction solution are as follows:

[0032] P1: 5'-ACATCGCCGGACAGGATGCTTCGGAGTACC-3';

[0033] P2: 3'-CACCCGCTGTTGTCTCACGACCTATACCGG-5'.

[0034] The PCR reaction system is 20 μL, including 15 μL of the PCR reaction solution, 2 μL of the primers and 3 μL of sample DNA;

[0035] The PCR reaction cycle parameters are:

[0036] 94°C, 2min; enter the cycle stage: denaturation at 94°C for 15s, annealing at 55°C for 1min, extension at 72°C for 20s, same as 35 cycles; stop at 10°C.

Embodiment 3

[0038] A human adenovirus universal nucleic acid detection kit, which includes an extraction solution for extracting human adenovirus DNA, a PCR reaction solution, and the reaction solution includes 50 μL of 13 μM Taq DNA polymerase, 100 μL of 150 μM dNTP, 200 μL of SyBr dye, and 15 μM of uracil glycosyl Lyase 50 μL, human adenovirus positive control substance, negative control substance;

[0039] The primer sequences used in the DNA amplification reaction in the PCR reaction solution are as follows:

[0040] P1: 5'-ACATCGCCGGACAGGATGCTTCGGAGTACC-3';

[0041] P2: 3'-CACCCGCTGTTGTCTCACGACCTATACCGG-5'.

[0042] The PCR reaction system is 20 μL, including 15 μL of the PCR reaction solution, 2 μL of the primers and 3 μL of sample DNA;

[0043] The PCR reaction cycle parameters are:

[0044] 94°C, 2min; enter the cycle stage: denature at 94°C for 15s, anneal at 60°C for 1min, extend at 72°C for 20s, same as 35 cycles; stop at 10°C.

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Abstract

The invention discloses a universal nucleic acid detection kit for human adenoviruses. The kit comprises an extracting solution for extracting human adenovirus DNA and a PCR reaction solution, and thereaction solution comprises 50 [mu] L of 13 [mu] M Taq DNA polymerase, 100 [mu] L of 150 [mu] M dNTP, 200 [mu] L of SyBr dye, and 50 [mu] L of 15 [mu] M uracil glycosylase. A human adenovirus positive reference substance and a human adenovirus negative reference substance; wherein the PCR reaction system is 20 [mu] L and comprises 15 [mu] L of a PCR reaction solution, 2 [mu] L of a primer and 3 [mu] L of sample DNA, and the sequence of the primer for DNA amplification reaction in the PCR reaction solution is as follows: P1: 5 '-ACATGAGGATGCTGAGTACC-3'; and P2: 3 '-CACCCCGCTGTTGTCTCTCTACCTACACCGG-5'. The universal nucleic acid detection kit and method for the human adenoviruses have very good specificity, can detect the human adenoviruses, are mature, quick and accurate in detection method, and provide reliable experimental basis for detection of the human adenoviruses.

Description

technical field [0001] The invention belongs to the technical field of virus detection, and in particular relates to a human adenovirus universal nucleic acid detection kit and method. Background technique [0002] Human adenoviruses (HAdV) belong to the Adenoviridae family and are a group of widely distributed DNA viruses, which can be divided into six subspecies A to F, with a total of 51 serotypes. Adenovirus is a non-enveloped virus, which can exist stably in a low pH environment and has strong resistance to physical and chemical agents. Adenovirus can produce tolerance to gastrointestinal secretions and bile, so adenovirus can Replicates in the gastrointestinal tract, producing high viral loads. Adenoviruses often multiply in the pharynx, conjunctiva, intestinal tract, and lymphoid tissues, and cause a variety of clinical symptoms, such as respiratory infections, conjunctivitis, gastroenteritis, hepatitis, hemorrhagic cystitis, and nervous system disorders Wait. Diff...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6869C12R1/93
CPCC12Q1/701C12Q1/6869C12Q2531/113C12Q2565/125
Inventor 金勇丰张鹏谢珍邢宽何志健
Owner 嘉兴实践医学科技有限公司
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