Separated nucleotide molecule and application thereof in bunia virus detection

A bunyavirus and nucleotide technology, which is applied in the detection/testing of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc., can solve the problems of lack of diagnosis and detection methods for diseases, unfavorable prevention and control of viral infection, etc. To achieve the effect of ensuring accuracy and feasibility, increasing the amplification factor, and benefiting the sensitivity

Inactive Publication Date: 2013-11-27
ICDC CHINA CDC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, clinicians, staff of CDCs at all levels and technicians in the field of microbiology still lack a basic understanding of the virus, and lack of diagnostic and detection methods for diseases caused by this virus infection, as well as differential diagnosis from other microbial infections. information, which is extremely detrimental to the prevention and control of the virus infection

Method used

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  • Separated nucleotide molecule and application thereof in bunia virus detection
  • Separated nucleotide molecule and application thereof in bunia virus detection
  • Separated nucleotide molecule and application thereof in bunia virus detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Embodiment 1 nested RT-PCR detection method

[0055] (1) Nested RT-PCR primers and primer sequences involved in the present invention.

[0056] 1. The first set of nested RR-PCR primers and primer sequences:

[0057] SEQ ID NO: 1: GTGGAGAATGGGCTGTCTCTT;

[0058] SEQ ID NO: 2: GGCCTGTACGACCAAATCTAC;

[0059] SEQ ID NO: 3: CCACATTGCCTAGCGAGACAT;

[0060] SEQ ID NO: 4: AGCCTGAGTCGGTCTTGATGT.

[0061] SEQ ID NO: 1-4 are respectively located at 50-70, 80-100, 1128-1148, 1050-1070 nucleotide positions of SEQ ID NO: 25.

[0062] The first round of RT-PCR uses primers SEQ ID NO: 1 and 3, and the expected size of the product is 1099bp;

[0063] The second round of nested PCR used primers SEQ ID NO: 2 and 4, and the expected size of the product was 991 bp.

[0064] 2. The second set of nested RT-PCR primers and primer sequences:

[0065] SEQ ID NO: 5: GGGCAAAGTGCCTTCTAGATG;

[0066] SEQ ID NO: 6: CATTGCCATCTCTGATGATCC;

[0067] SEQ ID NO: 7: CTTAGACTTGTCCTCCAGACT;

[00...

Embodiment 2

[0154] Example 2 Sample hybridization detection (Dot blot detection)

[0155] (1) main reagent involved in the experiment of the present invention

[0156] 1. Reverse transcriptase PrimeScript RTase and Buffer ( One StepRT-PCR Kit Ver.2), purchased from Bao Biological Engineering (Dalian) Co., Ltd.

[0157] 2. DNA polymerase La Taq DNA polymerase and Buffer were purchased from Treasure Bioengineering (Dalian) Co., Ltd.

[0158] 3. dNTPS was purchased from Bao Biological Engineering (Dalian) Co., Ltd.

[0159] 4. DNase and RNase-free ultrapure water, UltraPure TM DNase / RNase-Free Distilled Water was purchased from Invitrogen, USA.

[0160] 5. The sample RNA extraction kit, QIAamp MinElute Virus Spin Kit, was purchased from Qiagen, Germany.

[0161] 6. PCR product gel extraction kit, QIAquick Gel Extraction Kit, purchased from Qiagen, Germany.

[0162] 7. Nucleic acid hybrid nylon membrane (Hybond N+ membrane), purchased from GE Company of the United States.

[0163] 8....

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Abstract

The invention relates to a separated nucleotide molecule, application of the nucleotide molecule in bunia virus detection and a kit for the bunia virus detection. By the aid of a nucleotide sequence, a detection method and the kit, bunia viruses can be detected quickly, peculiarly and sensitively.

Description

technical field [0001] The invention relates to an isolated nucleotide molecule and its application in bunyavirus detection, as well as a bunyavirus detection kit. Background technique [0002] The family Bunyaviridae has more than 300 virus members divided into 5 genera, including Orthobunyavirus, Hantavirus, Nairovirus, Sandfly virus Phlebovirus and Tospovirus. [0003] Bunyakovirus particles are generally spherical with an envelope, about 80-120 nm in diameter, with 5-10 nm-long glycoprotein protrusions on the surface, and these protrusions are embedded in a double-layer lipid membrane with a thickness of about 5 nm. All members of this family of viruses have three-segment genomes, which are large (Large, L), medium (Medium, M), and small (Small, S) 3 antisense or ambisense linear ssRNAs, encoding 4 structural proteins, including Two outer membrane glycoproteins (Gn / Gc), a nucleoprotein (N) and a very large RNA polymerase (L). The terminal nucleotides of each genomic R...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/11C12Q1/70C12Q1/68
Inventor 熊衍文罗雪莲孙强正李娟叶长芸张永振徐建国
Owner ICDC CHINA CDC
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