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Nucleotide sequence for fast inspecting HIV virus, its method and extra-diagnostic reagent kit

A nucleotide sequence and in vitro diagnostic technology, applied in the fields of medical biology and virus molecular biology, can solve problems such as expensive, underestimated, and unguaranteed changes in combination

Inactive Publication Date: 2007-04-04
卫生部北京医院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This reagent uses the principle of RT-PCR-ELISA for detection, and only uses a pair of primers and probes corresponding to the HIV gag gene, which cannot guarantee the specific binding to each subtype and the binding ability caused by possible mutations in the primer or probe binding regions. There are reports in the literature that the kit has missed detection or quantitative underestimation
In addition, the kit is expensive, and the detection cost is about 50-80 US dollars per person, which limits its popularization in China

Method used

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  • Nucleotide sequence for fast inspecting HIV virus, its method and extra-diagnostic reagent kit
  • Nucleotide sequence for fast inspecting HIV virus, its method and extra-diagnostic reagent kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Embodiment 1. detects the primer and probe sequence of HIV virus

[0069] Commissioned by Shanghai Jikang Biotechnology Co., Ltd. to synthesize

[0070] Primer 1 forward 5'-ACATCAAGCAGCCATGCAAAT-3' SEQ ID No.1

[0071] Primer 1 reverse 5'-CTATGTCACTTCCCCTTGGTTCTCT-3' SEQ ID No.2

[0072] Probe 1 5'-ACCATCAATGAGGAAGCTGCAGAATGGG-3' SEQ ID No.3

[0073] Primer 2 forward 5'-GCTTTCAGCCCAGAAGTAATACC-3' SEQ ID No.4

[0074] Primer 2 Reverse 5'-ATTTGCATAGCTGCTTGATGTCC-3' SEQ ID No.5

[0075] Probe 2 5'-TCAGCATTATCAGAAGGAGCCACCCCACA-3'SEQ ID No.6

[0076] Probe 3 5'-TCAGGCCCCCTCAAAGCCGAC-3' SEQ ID No.7

[0077] Among them, the 5' ends of probe 1 and probe 2 are labeled with the fluorescent reporter group FAM, and the 3' ends are respectively labeled with the fluorescent quencher group TAMRA; the 5' ends of probe 3 are labeled with the fluorescent reporter group VIC, and the 3' terminal fluorescent quencher TAMRA.

Embodiment 2

[0078] Example 2. In vitro diagnostic kit 1 for detecting HIV virus

[0079] 1. Composition

[0080] 1. RNA extraction reagent: one selected from Trizol or magnetic bead method extraction reagents, both containing 1000 copies / ml internal standard virus-like particles:

[0081] (1) Trizol: purchased from Invitrogen, containing 1000 copies / ml of internal standard virus-like particles, prepared by our laboratory, the method refers to open literature, and stored at 4°C.

[0082] (2) Magnetic bead method extraction reagent: including lysate and other reagents, purchased from Shanghai Kehua Bioengineering Co., Ltd.; its lysate contains 1000 copies / ml internal standard virus-like particles, prepared by our laboratory, method Refer to published literature and store at 4°C.

[0083] 2. Reaction solution: composed of 50mM Tris (use concentration 20mM), 125mM KCl (use concentration 50mM), 10mM MgCl 2 (use concentration 4mM), 500ug / ml BSA (use concentration 200ug / ml), 0.375mM Sorbitol ...

Embodiment 3

[0090] Example 3. In vitro diagnostic kit 2 for detecting HIV virus

[0091] 1. Composition

[0092] 1. RNA extraction reagent: one selected from Trizol or magnetic bead method extraction reagents, both containing 1000 copies / ml internal standard virus-like particles:

[0093] (1) Trizol: purchased from Invitrogen, containing 1000 copies / ml of internal standard virus-like particles, prepared by our laboratory, the method refers to open literature, and stored at 4°C.

[0094] (2) Magnetic bead method extraction reagent: including lysate and other reagents, purchased from Shanghai Kehua Bioengineering Co., Ltd.; its lysate contains 1000 copies / ml internal standard virus-like particles, prepared by our laboratory, method Refer to published literature and store at 4°C.

[0095] 2. Reaction solution: 50mM Tris (purchased from Sigma, US, concentration 20mM), 125mM KCl (purchased from Sigma, USA, concentration 50mM), 10mM MgCl 2 (purchased from U.S. Sigma Company, use concentratio...

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Abstract

A nucleotide sequence for inspecting HIV virus rapidly, its method and in-vitro diagnostic reagent knit are disclosed. It's cheap, simple and accurate and has wide inspection range, and it can be used to inspect A-H sub-family and O-family of M family.

Description

technical field [0001] The invention relates to a nucleotide sequence, a method and an in vitro diagnostic kit for rapid detection of HIV virus, belonging to the field of virus molecular biology and medical biotechnology. Background technique [0002] HIV is a human immunodeficiency virus group in the genus Lentivirus of the Retroviridae family. So far, according to serological response and viral nucleic acid sequence determination, the HIV prevalence in the world can be divided into 2 types: HIV-1 type and HIV-2 type. Within the HIV-1 type, according to the homology of the env gene encoding the envelope protein and the gag gene encoding the capsid protein, it is further divided into three groups: M group (main group), O group (peripheral group) and N group. Group (new group or non-M non-O group), M group can be divided into A-J10 subtypes. Between HIV-1 and HIV-2, the nucleotide sequence has 45% homology, and there is immune cross-reaction. The early diagnosis of HIV is ...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/68
Inventor 李金明黄杰王露楠邓巍杨昌梅孟双申子瑜
Owner 卫生部北京医院
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