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Method and kit for detecting deletion mutation of cell apoptosis regulator gene (BIM)

A detection method and gene deletion technology, which is applied in the fields of molecular biology and medicine, can solve the problems of time-consuming, high equipment requirements, and high false negative rate, and achieve the effects of reducing the possibility of pollution, simple method, and intuitive results

Inactive Publication Date: 2015-04-29
SHANGHAI CHEST HOSPITAL
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  • Description
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  • Application Information

AI Technical Summary

Problems solved by technology

Among them, the sequencing method is currently considered as the gold standard method for detecting gene mutations because it can read the change of each base of DNA, but because of the defect of low sensitivity of this method, when detecting clinical tumor tissue samples with high heterogeneity The false negative rate is high, and at the same time, there are still defects such as cumbersome sequencing steps, long time consumption, and high requirements for equipment and operators. Therefore, it is not easy to form clinical diagnostic products with standardized operations
In addition, methods such as DHPLC, gene chips, and liquid phase chips also have defects such as complex operations and high requirements for equipment. Therefore, they are currently only used in laboratories of scientific research units.

Method used

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  • Method and kit for detecting deletion mutation of cell apoptosis regulator gene (BIM)
  • Method and kit for detecting deletion mutation of cell apoptosis regulator gene (BIM)
  • Method and kit for detecting deletion mutation of cell apoptosis regulator gene (BIM)

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Experimental program
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Effect test

Embodiment 1

[0034] Embodiment 1 real-time fluorescent PCR detection

[0035] (1) Experimental materials

[0036] The 480II fluorescent quantitative PCR instrument was purchased from Roche Company of Switzerland, and the polymerase chain reaction solution (FastStart Universal SYBR Green Master) was purchased from Roche Company of Switzerland.

[0037] (2) Primer design and synthesis:

[0038] Using the partial sequence of the BIM gene as a template, the Primer Premier5 software was used to analyze and design primers, which were synthesized by Sangon Biotech.

[0039] Primers for detection:

[0040] BIM gene deletion detection upstream primer sequence:

[0041] 5'-ATACCATCCAGCTCTGTCTTCATAG-3' (SEQ ID NO 2)

[0042] BIM gene deletion detection downstream primer 1 sequence: 5'-CCCAACCTCTGACAAGTGACC' (SEQ ID NO 3).

[0043] BIM gene deletion detection downstream primer 2 sequence: 5'-TTGGTGGGAATGTAAAATGGC-3' (SEQ ID NO 4).

[0044] (3) Sample testing:

[0045] A total of 30 cases of l...

Embodiment 2

[0056] The 2903 bp deletion in the intron between the third and fourth exons of the BIM gene was detected by common PCR method, and 10 cases of the above-mentioned case samples and 10 control samples were selected for sequencing to determine the deletion of BIM.

[0057] (1) Experimental method

[0058] The PCR sequencing primers used above-mentioned fluorescent PCR primers were used, and the amplified products were subjected to agarose electrophoresis. The electrophoresis instrument was from BidRad Company of the United States.

[0059] (2) Experimental results

[0060] The screenshot of the result is as follows Figure 4 as shown,

[0061] In the end, the electrophoresis result of case X was the same as The analysis results of 480II fluorescent PCR were completely consistent.

[0062] (3) Association analysis of BIM gene 2903bp deletion and lung cancer susceptibility

[0063] The comparison of the distribution of BIM gene 2903bp deletion in lung cancer patients and con...

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Abstract

The invention belongs to the fields of molecular biology and medicine and relates to a method for detecting deletion mutation of a cell apoptosis regulator gene (BIM) and a detection kit applying the method. The method for detecting deletion mutation of the cell apoptosis regulator gene (BIM) comprises a step of detecting deletion of 2903bp on an intron between a third exon and a fourth exon on the BIM gene. The invention also discloses a corresponding detection kit which also contains a primer used for amplifying a commonly deleted region of 2903bp on the BIM. When the method and kit provided by the invention are utilized for detecting BIM deletion condition, operation is simple and easy, rapidness and high efficiency are realized, and cost is low.

Description

technical field [0001] The invention belongs to the fields of molecular biology and medicine, and relates to a detection method for the deletion mutation of the apoptosis mediator gene (BIM) and a detection kit thereof, in particular to a method for detecting the gap between the third exon and the fourth exon of the BIM gene The detection method and detection kit for the 2903bp deletion mutation on the intron. Background technique [0002] The prior art discloses that the BIM gene is a member of the BCL2 protein family encoding the apoptosis gene, and plays an important regulatory role in cell apoptosis. Studies have shown that the 2903bp deletion between exon 3 and exon 4 of BIM gene will cause the lack of expression of BIM subtype BH3 structure. This lack of expression can lead to greater drug resistance in lung cancer patients to some therapeutic drugs. Studies have shown that in cancer patients, this deletion is associated with the duration of drug response and can be ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 韩宝惠夏金晶邵敏华黄迅威
Owner SHANGHAI CHEST HOSPITAL
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