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Combined detection method and diagnostic kit of fusion genes related to lymphoma

A diagnostic kit and fusion gene technology, applied in the direction of microbial determination/examination, fluorescence/phosphorescence, biochemical equipment and methods, etc., can solve the problems that cannot truly meet clinical diagnostic testing, complex operation of fluorescence in situ hybridization technology, and insufficient sensitivity. Good and other problems, to achieve the effect of rapid detection, high specificity and high sensitivity

Active Publication Date: 2013-06-26
MEDI GENETECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Among the currently widely used molecular biology methods for detecting lymphoma fusion genes at home and abroad, fluorescence in situ hybridization (FISH) can only perform qualitative detection, and the operation is complicated; fluorescent quantitative PCR has limitations in detection throughput, so it is still Can not 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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  • Combined detection method and diagnostic kit of fusion genes related to lymphoma
  • Combined detection method and diagnostic kit of fusion genes related to lymphoma
  • Combined detection method and diagnostic kit of fusion genes related to lymphoma

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Example 1: A liquid-phase chip detection method for a lymphoma-related fusion gene

[0068] The specific detection method includes the following steps:

[0069] 1. Preparation of microsphere mixture for detection of API2-MALT1(A1446-M1123) fusion gene

[0070] 1. Synthesize oligonucleotide probes according to the following sequence:

[0071] API2-MALT1 (A1446-M1123): 5'-AminolinkerC12 CCAAGATTATTTAATTCATTTG-3', as shown in SEQ ID NO.2;

[0072] β-actin gene: 5'-AminolinkerC12 TCATTGTAGAAGGTGTGGTG-3', as shown in SEQ ID NO.15;

[0073] 2. Coupling the oligonucleotide probe containing amino modification with two kinds of carboxyl microspheres numbered 25 and 64 respectively

[0074] 2.1 Take out a small portion of fresh dry powdered EDC stored at -20°C and equilibrate to room temperature;

[0075] 2.2 with dH 2 O dissolve the oligonucleotide probes of API2-MALT1 (A1446-M1123) and β-actin respectively at a concentration of 1 mM (1 nmol / μl);

[0076] 2.3 Vortex the st...

Embodiment 2

[0169] Example 2: Liquid-phase chip combined detection method for two lymphoma-related fusion genes

[0170] The specific detection method includes the following steps:

[0171] 1. Preparation of microsphere mixture for detection of API2-MALT1 (A1446-M1123), NPM-ALK fusion gene

[0172] 1. Synthesize oligonucleotide probes according to the following sequence:

[0173] API2-MALT1 (A1446-M1123): 5'-AminolinkerC12 CCAAGATTATTTAATTCATTTG-3', as shown in SEQ ID NO.2;

[0174] NPM-ALK: 5'-AminolinkerC12 CCTATAGTTGTTTTAAATGC-3', as shown in SEQ ID NO.4;

[0175] β-actin gene: 5'-AminolinkerC12 TCATTGTAGAAGGTGTGGTG-3', as shown in SEQ ID NO.15;

[0176] 2. Coupling the oligonucleotide probes containing amino modifications to three kinds of carboxyl microspheres numbered 25, 50, and 64 respectively

[0177] 2.1 Take out a small portion of fresh dry powdered EDC stored at -20°C and equilibrate to room temperature;

[0178] 2.2 with dH 2 O dissolve the oligonucleotide probes of API...

Embodiment 3

[0273] Example 3: Liquid-phase chip combined detection method for 3 lymphoma-related fusion genes

[0274] The specific detection method includes the following steps:

[0275] 1. Preparation of microsphere mixture for detection of API2-MALT1 (A1446-M814), API2-MALT1 (A1446-M1123), and NPM-ALK fusion genes

[0276] 1. Synthesize oligonucleotide probes according to the following sequence:

[0277] API2-MALT1 (A1446-M814): 5'-AminolinkerC12 GCTTTGATTCTTTTTTCTCAG-3', as shown in SEQ ID NO.1;

[0278] API2-MALT1 (A1446-M1123): 5'-AminolinkerC12 CCAAGATTATTTAATTCATTTG-3', as shown in SEQ ID NO.2;

[0279] NPM-ALK: 5'-AminolinkerC12 CCTATAGTTGTTTTAAATGC-3', as shown in SEQ ID NO.4;

[0280] β-actin gene: 5'-AminolinkerC12 TCATTGTAGAAGGTGTGGTG-3', as shown in SEQ ID NO.15;

[0281] 2. Coupling the oligonucleotide probes containing amino modifications to four kinds of carboxyl microspheres numbered 11, 25, 50, and 64 respectively

[0282] 2.1 Take out a small portion of fresh dry ...

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Abstract

The invention discloses joint detection methods for lymphoma fusion genes and diagnostic kits thereof. After primers and probes for the mRNAs of lymphoma fusion genes API2-MALT1 (A1446-M814), API2-MALT1 (A1446-M1123), API2-MALT1 (A1446-M1150), NPM-ALK are designed, the probes are covalently bonded to the beads to form probe-bead mixtures, which are hybridized with the RT-PCR amplification products. After streptavidin-phycoerythrin is added, fluorescent signals from different beads can be detected, and it can be determined whether or not the sample to be detected contains the lymphoma fusion genes, and the expression profiles of the fusion genes can be determined.

Description

technical field [0001] The invention relates to the technical field of in vitro diagnosis and detection, in particular to a combined liquid phase chip detection method and a diagnostic kit for multiple lymphoma-related fusion genes. Background technique [0002] Lymphoma is a group of malignant tumors originating from lymph nodes or other lymphoid tissues. It can be divided into two categories: Hodgkin's disease (HD) and non-Hodgkin's lymphoma (NHL). one. Histologically, tumorous proliferation of lymphocytes and (or) histiocytes can be seen, and the main clinical manifestation is painless and progressive lymphadenopathy. [0003] Mucosa-associated lymphoid tissue (MALT) lymphoma is a malignant lymphoma that originates outside lymph nodes, and its incidence accounts for about 40% of all lymphomas, among which MALT lymphoma of the gastrointestinal tract is the most common. API2-MALT1 is a fusion gene produced by chromosomal translocation in MALT lymphoma, which can be found ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68G01N21/64
CPCC12Q2600/16C12Q1/6886C12Q2600/158
Inventor 邵棠袁福美
Owner MEDI GENETECH