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Kit for detecting NUP214-ABL1 gene relative expression quantity and method

A NUP214-ABL1, relative expression technology, applied in biochemical equipment and methods, recombinant DNA technology, DNA / RNA fragments, etc., can solve the problems of various reagents, tedious test process, time-consuming and laborious, etc., and achieve amplification efficiency and rate the best results

Inactive Publication Date: 2017-09-12
WUHAN ADICON CLINICAL LAB
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

FISH test results are relatively intuitive, but the test process is cumbersome, involving a wide variety of reagents, time-consuming and labor-intensive, and the results need to be interpreted by experienced professionals, and the interpretation of the results is relatively subjective

Method used

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  • Kit for detecting NUP214-ABL1 gene relative expression quantity and method
  • Kit for detecting NUP214-ABL1 gene relative expression quantity and method
  • Kit for detecting NUP214-ABL1 gene relative expression quantity and method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] The present invention is used for assisting the formulation of clinical individualized treatment plan for acute T lymphocytic leukemia (T-cell acute lymphoblasticleukemia, T-ALL). Mainly include the following reagents: erythrocyte lysate; TRIzol; chloroform; isopropanol; absolute ethanol;

[0045] Detection system PCR reaction solution: ReverTra Ace qPCR RT Kit (TOYOBO); THUNDERBIRDProbe qPCR Mix (2×), ABL internal reference gene and NUP214-ABL1 target gene primers and probes are all 10 μM;

[0046] The primers and probes for detecting the internal reference gene ABL and the target gene NUP214-ABL1 are respectively:

[0047]

[0048] Positive control substance: solution containing NUP214-ABL1 genome;

[0049] Negative control substance: solution without NUP214-ABL1 genome.

Embodiment 2

[0051] The operating flow of the inventive method:

[0052] (1) Extraction of total RNA in blood: Add 1ml of erythrocyte lysate into a clean 1.5ml centrifuge tube, take 0.5ml of anticoagulated blood and mix well. Let stand at room temperature for 10 minutes; centrifuge at 1500rpm for 5min, discard the supernatant, and collect the cells at the bottom; add 0.5ml red blood cell lysate again, centrifuge at 1500rpm for 5min, discard the supernatant, and collect the cells at the bottom; add 1ml TRIzol to the cells, and pipette repeatedly until the precipitation is complete Dissolve, let stand at room temperature for 5 minutes; add 0.2ml chloroform, oscillate evenly; centrifuge at 14000rpm at 4°C for 10 minutes, absorb the supernatant layer and transfer to another new centrifuge tube; add an equal volume of isopropanol, mix well up and down, and let stand at room temperature for 10 minutes Centrifuge at 14000rpm at 4°C for 10min, discard the supernatant, add 1ml of 75% ethanol, wash ...

Embodiment 3

[0062] Using the nucleic acid detection method of the present invention to detect samples from healthy people

[0063] Take 12 cases of healthy physical examination samples to be tested, extract the genome, prepare reagents and detect according to the method described in Example 2.

[0064] Add 2 μL of each sample to the detection system PCR reaction solution. At the same time, make a standard curve of positive, negative, blank control, and internal reference gene / target gene. A 96-well fluorescent PCR instrument can detect 12 samples at the same time, each sample has 2 repetitions, one positive control and one negative control, and the detection time is only 100 minutes. The abl of all samples in the 12 screening samples had a line, but none of the samples of NUP214-ABL1 had a line. The results are shown in Table 1 and Figure 1-5 shown

[0065] Table 1 NUP214-ABL1 mRNA expression levels in 12 healthy physical examination samples

[0066]

[0067]

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Abstract

The invention provides a kit for detecting the NUP214-ABL1 gene relative expression quantity. A real-time fluorescence PCR technology is used for detecting NUP214-ABL1 fusion gene. The kit comprises PCR reaction liquid, wherein the PCR reaction liquid comprises a probe and a primer for amplifying the NUP214-ABL1 fusion gene. Through the test, the kit has good specificity and high flexibility; the operation is simple and convenient. The detection of the NUP214-ABL1 fusion gene in a patient with acute T lymphocytic leukemia in clinics is facilitated; the important significance is realized on treatment scheme regulation, treatment effect evaluation, prognosis prediction and clinic recurrence prevention.

Description

technical field [0001] The invention belongs to the field of disease gene detection, and in particular relates to a kit and a method for detecting the expression of NUP214-ABL1 in human acute T lymphocytic leukemia by using probe real-time fluorescent PCR technology. technical background [0002] Acute lymphoblastic leukemia (T-cell acute lymphoblastic leukemia, T-ALL) is a hematological tumor characterized by malignant clonal proliferation of T lymphocytes. 25% or so. T-ALL often occurs in children and young adults. The main features of patients are abnormally high peripheral blood leukocyte counts and high proportion of blast cells. They are prone to T lymphocyte bone marrow infiltration, pleural effusion, central nervous system infiltration, and mediastinal tumors. Complications, dangerous condition, poor prognosis. [0003] Previous studies have found that more than 50% of T-ALL patients can detect abnormal karyotype, and cytogenetic testing has become an important dia...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6886C12Q1/686C12Q2600/158C12Q2561/113C12Q2563/107C12Q2561/101
Inventor 牛林梅吴鹏飞王淑一
Owner WUHAN ADICON CLINICAL LAB
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