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Leukemia fusion gene combined parallel detecting method and diagnostic reagent kit

A technology that integrates genes and detection methods is applied in the measurement/testing of microorganisms, biochemical equipment and methods, fluorescence/phosphorescence, etc. It can solve the problems of cumbersome operation, poor repeatability, and complicated operation of fluorescence in situ hybridization technology, and achieve High sensitivity and high specificity effect

Inactive Publication Date: 2010-09-22
MEDI GENETECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Immunological methods have high false positive and false negative rates, and cannot achieve early diagnosis
Among the molecular biology methods widely used at home and abroad to detect leukemia fusion genes, fluorescence in situ hybridization (FISH) can only perform qualitative detection, and the operation is complicated; fluorescent quantitative PCR has limitations in detection throughput, so none of them can Really meet the needs of clinical diagnostic testing
The traditional solid-phase biochip (Biochip) technology has outstanding weaknesses such as poor repeatability, insufficient sensitivity, and cumbersome operations.

Method used

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  • Leukemia fusion gene combined parallel detecting method and diagnostic reagent kit
  • Leukemia fusion gene combined parallel detecting method and diagnostic reagent kit
  • Leukemia fusion gene combined parallel detecting method and diagnostic reagent kit

Examples

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Effect test

example 1

[0078] Example 1: Liquid-phase chip joint parallel detection method for four leukemia fusion genes

[0079] The specific detection method includes the following steps:

[0080] 1. Preparation of microsphere mixture for detecting four fusion genes of BCR-ABL (b2a2), TEL-AML1, CBFB-MYH11 (A type), and PML-RARA (L form)

[0081] 1. Synthesize oligonucleotide probes according to the following sequence:

[0082] BCR-ABL(b2a2)5'-AminolinkerC12TGAAGGGCTTCTTCCTTATT-3'

[0083] TEL-AML 15'-AminolinkerC12TCCAAGTATGCATTCTGCTA-3'

[0084] CBFB-MYH11(A type)5'-AminolinkerC12GCTCATGGACCTCCATTTCC-3'

[0085] PML-RARA(L form)5'-AminolinkerC12GTCTCAATGGCTGCCTCCCC-3'

[0086] Abelson gene 5'-AminolinkerC12CTGAAGGGCTTCTTCCAGAT-3'

[0087] 2. Coupling oligonucleotide probes containing amino modifications to five carboxyl microspheres numbered 11, 25, 37, 50, and 64

[0088] 2.1 Take out a small portion of fresh dry powdered EDC stored at -20°C and equilibrate to room temperature;

[0089] ...

example 2

[0194] Example 2: Liquid-phase chip joint parallel detection method for 10 kinds of leukemia fusion genes

[0195] The specific detection method includes the following steps:

[0196] 1. Detection of BCR-ABL(b2a2), BCR-ABL(b3a2), BCR-ABL(e1a2), E2A-PBX1, TEL-AML1, MLL-AF4(e10 / e4), CBFB-MYH11(A type), AML1 - Preparation of microsphere mixture of ten fusion genes of ETO, PML-RARA(L form), PML-RARA(Sform)

[0197] 1. Synthesize oligonucleotide probes according to the following sequence:

[0198] BCR-ABL(b2a2) 5'-AminolinkerC12TGAAGGGCTTCTTCCTTATT-3'

[0199] BCR-ABL(b3a2) 5'-AminolinkerC12TGAAGGGCTTTTGAACTCTG-3'

[0200] BCR-ABL(e1a2) 5'-AminolinkerC12TGAAGGGCTTCTGCGTCTCC-3'

[0201] E2A-PBX 1 5’-AminolinkerC12TACTCAAAACACTGTAGGAG-3’

[0202] TEL-AML 1 5'-AminolinkerC12TCCAAGTATGCATTCTGCTA-3'

[0203] MLL-AF4(e10 / e4) 5'-AminolinkerC12AGTAGGTCTGCTTAAAGTCC-3'

[0204] CBFB-MYH11(A type)5'-AminolinkerC12GCTCATGGACCTCCATTTCC-3'

[0205] AML1-ETO 5'-AminolinkerC12TCAGTACGATTTC...

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Abstract

The invention discloses a leukemia fusion gene combined parallel detecting method and a diagnostic reagent kit. The combined parallel detecting method is characterized in that multiple fusion genes related to leukemia can be detected at one time. By designing a primer and a probe of the leukemia fusion gene mRNA, the mixture covalently combined by the probe and microspheres is hybridized with a reverse transcription PCR amplified product, the streptavidin of phycoerythrin is added, and then the fluorescence signals of different microspheres can be detected, thereby determining whether the sample to be detected contains leukemia fusion genes or not and the express conditions of the fusion genes. The diagnostic reagent kit has the advantages of high sensitivity, high flux property, quick and accurate detection, and the like, and can simultaneously carry out qualitative and quantitative detection on multiple leukemia fusion genes.

Description

technical field [0001] The invention relates to the technical field of in vitro diagnosis and detection, in particular to a liquid-phase chip combined parallel detection method for multiple leukemia fusion genes and a diagnostic kit thereof. Background technique [0002] Leukemia is a malignant tumor of the hematopoietic system caused by abnormal differentiation and malignant proliferation of hematopoietic stem cells. It is one of the most common malignant tumors in my country. With the continuous progress of leukemia research, it has been found that many leukemia patients have specific chromosomal translocations, leading to the generation of new fusion genes, and these abnormal genes have become molecular biology-specific markers of different types of leukemia. In 2000, WHO's criteria for leukemia classification included some common abnormal genes as the criteria for the basic diagnosis of leukemia. Therefore, the detection of leukemia-related fusion genes at the molecular...

Claims

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Application Information

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IPC IPC(8): C12Q1/68G01N21/64
Inventor 邵棠孙黎徐春雷
Owner MEDI GENETECH
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