Hypersensitivity Epstein-Barr (EB) virus fluorescence quantitative polymerase chain reaction (PCR) kit for locked nucleotide acid (LNA) and detection method and application thereof

A fluorescence quantitative, EB virus technology, applied in the field of molecular biology, to achieve the effect of short detection time, reduced result deviation, and large sample size

Inactive Publication Date: 2010-12-01
广州达健生物科技有限公司
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0007] Most of the above methods were compared with traditional methods at the laboratory level, but only few studies evaluated their performance in actual clinical and applied domains

Method used

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  • Hypersensitivity Epstein-Barr (EB) virus fluorescence quantitative polymerase chain reaction (PCR) kit for locked nucleotide acid (LNA) and detection method and application thereof
  • Hypersensitivity Epstein-Barr (EB) virus fluorescence quantitative polymerase chain reaction (PCR) kit for locked nucleotide acid (LNA) and detection method and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0038] Example 1: Reproducibility experiment of fluorescent quantitative PCR kit detecting EBV-DNA in plasma of patients with nasopharyngeal carcinoma

[0039] The Epstein-Barr virus fluorescence quantitative PCR kit of locked nucleotide (LNA) sensitization of the present invention comprises LNA (locked nucleotide)-TaqMan fluorescence quantitative reaction solution, positive control sample, wherein, LNA-TaqMan fluorescence quantitative reaction solution comprises:

[0040] Forward primer: 5'-AAT TTT TTC TGC TAA GCC CAA CA-3';

[0041] Reverse primer: 5'-ACG GGT GGG TGT GTG TAG TGT-3';

[0042] LNA-TanMan fluorescent probe: 5'-FAM-CCA CCA CAC CCA GGC-MGB3'.

[0043] The positive control sample is the plasmid DNA containing the Epstein-Barr virus genome fragment.

[0044] To use this kit to detect EBV-DNA, proceed as follows:

[0045] (1) Preparation of fluorescent quantitative nucleic acid amplification reaction system: add 0.15 μl (20 μM) of forward primer, 0.15 μl (20 μM) ...

Embodiment 2

[0049] Embodiment 2: The EBV-DNA fluorescent quantitative PCR kit of the present invention detects and detects the plasma EBV-DNA concentration of nasopharyngeal carcinoma patients

[0050] Fig. 2 is a preferred result diagram of detecting plasma EBV-DNA concentration of nasopharyngeal carcinoma patients with the EBV-DNA fluorescent quantitative PCR kit of the present invention, the method is the same as that described in Example 1. in, Figure 2A It shows that the sample amplification curve is clear, and the negative control has no amplification; Figure 2B Standard curve for comparison. The results of PCR amplification were analyzed automatically with the automatic analysis software SDS (Version 2.0). The quantitative results of plasma EBV-DNA were converted according to the formula: C=Q×(V DNA / V PCR )×(1 / V EXT ). Wherein C is the concentration (copy / ml) of EBV-DNA in the plasma to be tested, Q is the EBV-DNA concentration that PCR reaction obtains, V DNA Final dilut...

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Abstract

The invention provides a hypersensitivity Epstein-Barr (EB) virus fluorescence quantitative polymerase chain reaction (PCR) kit for locked nucleotide acid (LNA). The kit comprises LNA-TaqMan reaction liquid, a positive control sample, a forward primer, a reverse primer, and an LNA-TanMan fluorescent probe, wherein the forward primer is 5'-AATTTTTTCTGCTAAGCCCAACA-3'; the reverse primer is 5'-ACGGGTGGGTGTGTGTAGTGT-3'; and the LNA-TanMan fluorescent probe is 5'-FAM-CCACCACACCCAGGC-MGB3'. The invention also provides a hypersensitivity EB virus fluorescence quantitative PCR detection method for the LNA and the application of the kit in the detection of EB viruses, the diagnosis and treatment of nasopharyngeal darcinoma, prognosis judgment and the monitoring of recurrence and transfer after the treatment of the nasopharyngeal darcinoma. The kit of the invention has the advantages of rapidness, high sensitivity, simple operation process, large detection sample size and safety.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a locked nucleotide (LNA)-sensitized Epstein-Barr virus fluorescent quantitative PCR detection kit and its detection method and application, which are applicable to various tissues, cells, blood and other body fluids Quantitative detection of Epstein-Barr virus nucleic acid. Background technique [0002] Epstein-Barr Virus (EBV) is an important DNA virus that infects humans. EBV is a DNA virus, belonging to the herpesvirus γ subfamily, and herpes simplex virus type I (HSV-1) and type II (HSV-2), cytomegalovirus (CMV), varicella-herpes virus (VZV) and recently discovered The human herpesvirus type 6 belongs to the human herpesvirus group, and they are indistinguishable from each other in morphology, and their propagation modes are similar. The determination of the entire nucleotide sequence of the EBV genome has been completed. The genome is a double-stranded DNA molecule consi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68G01N21/64C12R1/93
Inventor 石大陆
Owner 广州达健生物科技有限公司
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