Kit for quantitatively detecting IKZF1 gene mutation

A quantitative detection and kit technology, which is applied in the determination/testing of microorganisms, DNA/RNA fragments, recombinant DNA technology, etc., can solve the problem of inability to quantitatively analyze the level of IKZF1 mutation

Inactive Publication Date: 2013-05-15
PEOPLES HOSPITAL PEKING UNIV
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

[0005] At present, the method of detecting IKZF1 mutation is mainly the met...

Method used

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  • Kit for quantitatively detecting IKZF1 gene mutation
  • Kit for quantitatively detecting IKZF1 gene mutation
  • Kit for quantitatively detecting IKZF1 gene mutation

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Embodiment 1, detection IKZF1 gene mutation

[0020] 1. Test samples

[0021] 306 cases of adult newly diagnosed ALL, 10 cases of non-hodgkin's lymphoma (NHL), 21 cases of chronic lymphocytic leukemia (chronic lymphoblastic leukemia, CLL), diagnostic criteria according to the new classification criteria of the World Health Organization. The samples for extracting DNA were the peripheral blood or bone marrow of the patients mentioned above, all of which were informed consent of the patients.

[0022] 2. Sequencing method to detect IKZF1 gene mutation

[0023] Using the extracted DNA as a template, the upstream primer: 5-ccacagggcaagtcatccacattttg-3' and the downstream primer: 5-cagaccatagagtccctcctaggggaaaaa-3' were used for PCR amplification. The PCR reaction system is 50 μl: 25 μl of 2×PCR amplification premix reagent (Tiangen Biological Co., Ltd.), 400 nM of upstream and downstream primers, 300 ng of DNA. The PCR reaction conditions are as follows: first 95°C for 5...

Embodiment 2

[0058] Embodiment 2, the sensitivity of the inventive method

[0059] 1. Detection sensitivity of plasmid standard:

[0060] The plasmid standard was diluted tenfold (10 7 , 10 6 , 10 5 , 10 4 , 10 3 , 10 2 , 10 1 , 10 0 copies), followed by real-time fluorescent quantitative PCR. Each gradient was repeated 3 times.

[0061] The results showed that the correlation coefficient was above 0.99, and the detection sensitivity could reach 1 copy. The amplification efficiencies of ALB and IKZF1 mutations were similar (standard curve slopes were -3.45 and -3.56, respectively).

[0062] 2. Detection sensitivity for IKZF1 mutation-positive patients:

[0063] The DNA samples of 8 IKZF1 mutation-positive patients were serially diluted 10 times (the initial concentration of DNA was 150ng / ul), and then real-time fluorescent quantitative PCR was performed.

[0064] As a result, a sensitivity of 10 -4 or 10 -5 , and the correlation coefficients are all above 0.99. The results...

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Abstract

The invention discloses a kit for quantitatively detecting IKZF1 gene mutation. The kit for quantitatively detecting the IKZF1 gene mutation consists of a primer pair and a probe, wherein a primer sequence of the primer pair is shown as SEQ ID No:1; the other primer sequence is shown as SEQ ID No:2; and the sequence of the probe is shown as SEQ ID No:3. The primer pair and the probe can be used for quantitatively detecting the IKZF1 gene mutation level, and the kit is easy, convenient and sensitive, so that the kit is used for auxiliary diagnosis, MRD (Minimal Residual Disease) monitoring, prognostic evaluation and the like on neoplastic hematologic disorder patients (particularly ALL leukemia patients and CML patients) and has significance in the field of medical detection.

Description

technical field [0001] The invention relates to a kit for quantitative detection of IKZF1 gene mutation. Background technique [0002] Ph-positive leukemia includes chronic myelogenous leukemia (chronic myelogenous leukemia, CML) and certain acute lymphoblastic leukemia (acute lymphoblastic leukemia, ALL), the instigator of which is the BCR-ABL1 fusion gene caused by the Ph chromosome. However, therapy targeting BCR-ABL1 has not been effective in some Ph-positive leukemias. The latest research has found that the role of other genetic factors including IKZF1 gene deletion cannot be ignored in the occurrence and progression of this type of leukemia. [0003] IKZF1 gene encodes a transcription factor protein with zinc finger structure, which plays an important regulatory role in the differentiation and development of lymphocytes. Ikaros, the downstream product of IKZF1 gene, is a lymphocyte-specific transcription factor. Deletion of the IKZF1 gene can result in underexpressi...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
Inventor 阮国瑞黄晓军江滨刘开彦陈珊珊江浩刘艳荣赖悦云李玲娣冷馨黎宁
Owner PEOPLES HOSPITAL PEKING UNIV
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