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Human cytomegalovirus real-time fluorescent nucleic acid constant temperature amplification detection kit

A human cytomegalovirus, real-time fluorescence technology, applied in the field of virus biological detection, achieves the effects of simple equipment, reduced design and production costs, and monitoring the existence of false negatives

Active Publication Date: 2019-10-11
SHANGHAI RENDU BIOTECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the problems faced by the application of SAT technology in the detection of different types of viruses are different, and it is necessary to specifically analyze the characteristics of the virus for special design.
At present, there is no research report on real-time fluorescent nucleic acid constant temperature amplification detection technology for human cytomegalovirus (HCMV) in China

Method used

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  • Human cytomegalovirus real-time fluorescent nucleic acid constant temperature amplification detection kit
  • Human cytomegalovirus real-time fluorescent nucleic acid constant temperature amplification detection kit
  • Human cytomegalovirus real-time fluorescent nucleic acid constant temperature amplification detection kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1 Design of special primers and probes for real-time fluorescent nucleic acid constant temperature amplification detection of human cytomegalovirus (HCMV)

[0047] The present invention selects a highly conserved and specific segment in the HCMV PP67 gene as the amplification target sequence region, and uses DNAStar, DNAMAN software and artificial design for real-time fluorescent nucleic acid constant temperature amplification to detect human giant cells according to the principle of primer probe design The specific primer and probe sequence of virus (HCMV) obtain following specific sequence:

[0048] (1) A capture probe (TCO, Target Capture Oligo) that can specifically bind to the target nucleic acid (HCMV RNA) sequence of human cytomegalovirus (HCMV), the nucleotide sequence of the capture probe is: 5' ACGTCACGCGATGCCTACTCCGATAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA 3';

[0049] (2) A pair of HCMV amplification primers for producing DNA copies of the HCMV target nucle...

Embodiment 2

[0055] Embodiment 2: The design of the reference substance used for real-time fluorescent nucleic acid constant temperature amplification detection of human cytomegalovirus (HCMV)

[0056] Because the SAT amplification is susceptible to the influence of various factors, the amplification fails, causing the kit user to make a wrong judgment and draw a wrong conclusion. Therefore, a control can also be set in the kit of the present invention to eliminate the distortion of the detection result. Condition.

[0057] The reference substances that can be set in the present invention include: one or more control substances in HCMV positive control, HCMV negative control and HCMV internal standard.

[0058] The HCMV positive control can be RNA transcribed in vitro, or human cytomegalovirus double-stranded DNA. By detecting the positive control, it can be proved that the detection method and materials of the kit are correct, and the accuracy of the detection can be guaranteed. At the s...

Embodiment 3

[0072] Example 3 Preparation of a real-time fluorescent nucleic acid constant temperature amplification detection kit for human cytomegalovirus (HCMV)

[0073] Using the special primers and probes provided in Example 1, a real-time fluorescent nucleic acid constant temperature amplification detection kit for human cytomegalovirus (HCMV) of the present invention was obtained. The kit contains components such as capture probe (TCO, Target Capture Oligo), T7 primer, nT7 primer, HCMV detection probe, M-MLV reverse transcriptase and T7 RNA polymerase, among which:

[0074] The nucleotide sequence of the capture probe is sequence 2, the sequence of the T7 primer is sequence 3, the sequence of the nT7 primer is sequence 4, the nucleotide sequence of the detection probe is sequence 5, and the nucleotide sequence of the internal standard probe is sequence 7.

[0075] The capture probe exists in the viral nucleic acid extraction solution, the T7 primer, nT7 primer and HCMV detection pro...

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Abstract

The invention discloses a realtime fluorescent nucleic acid constant temperature amplification detection kit of human cytomegalovirus (HCMV). The detection kit comprises a capture probe, a HCMVT7 primer, an nT7 primer, a HCMV detection probe, M-MLV reverse transcriptase and T7RNA polymerase. The provided method can high specifically, high sensitively, and rapidly carry out nucleic acid amplification detection on samples containing HCMV such as urine, milk, blood, and the like, and little pollution is generated. The provided kit and method can accurately, rapidly, and conveniently carrying out qualitative detection on HCMV; diagnose the virus infection, monitor and predict epidemicity of HCMV, helps the auxiliary diagnosis of virus infection, and observe the drug therapeutic effect.

Description

technical field [0001] The invention relates to the technical field of biological detection of viruses, in particular to primers and probes used in the real-time fluorescent nucleic acid constant temperature amplification detection of human cytomegalovirus (HCMV) by combining specific target capture technology and real-time fluorescent nucleic acid constant temperature amplification detection technology. Needles and related kits. Background technique [0002] Human cytomegalovirus (HCMV), also known as sialovirus or human herpesvirus 5, is the largest virus in the human herpesvirus group. Together with toxoplasma, rubella virus, herpes simplex virus and Treponema pallidum, it is called the five major biological teratogenic factors of human beings. After this virus enters the cell, it will cause the cell to enlarge, so it is called cytomegalovirus. Cytomegalovirus infection is widespread in my country. Normally, many normal people have CMV latent in the salivary glands, bre...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/6844C12R1/93
Inventor 冯俊居金良
Owner SHANGHAI RENDU BIOTECH
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