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Acute myelogenous leukemia miRNA marker

A technology for acute myeloid and leukemia, applied in the field of biomedicine, can solve problems such as abnormal gene expression and cancer induction

Active Publication Date: 2015-11-18
QINGDAO MEDINTELL BIOMEDICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The abnormal expression of miRNA directly leads to the abnormal expression of some genes related to the occurrence of diseases, and induces the occurrence of cancer.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Example 1 Screening of miRNAs associated with acute myeloid leukemia

[0049] 1. Sample acquisition: Collect blood samples from 10 healthy people and 10 patients with acute myeloid leukemia. All the above specimens were obtained with the consent of the organizational ethics committee.

[0050] 2. Extraction of total RNA from samples

[0051] Total RNA was extracted using the BloodRNA extraction kit from U-gene. Specific steps are as follows:

[0052] 1) Add 5 times the volume of 1×XR-I buffer per volume of fresh blood (maximum 1ml), for example: add 5ml of XR-I buffer per 1ml of blood, and vortex to mix;

[0053] 2) Ice-bathed for 15 minutes, quickly mixed twice on a vortex shaker, the solution became clear, indicating that the red blood cells had been lysed. If the hematocrit or ECR of individual samples increases, extend the ice bath time to 20 minutes;

[0054] 3) Centrifuge at 450g for 10min at 4°C to precipitate white blood cells, and completely discard the su...

Embodiment 2

[0080] Example 2 QPCR verification of differentially expressed miRNA-1262

[0081] 1. Select miRNA-1262 according to the detection results of the miRNA chip for large sample QPCR verification. According to the sample collection method in Example 1, 80 blood samples from patients with acute myeloid leukemia and 80 blood samples from healthy people were selected.

[0082] 2. The RNA extraction process is the same as in Example 1.

[0083] 3. Reverse transcription:

[0084] 1) Mix 10 pg-1 μg of total RNA template with 2 μl 10× buffer, 2 μl ldATP (10 mM), 0.5 μl polyA polymerase, 0.5 μl ribonuclease (RNase) inhibitor and ribonuclease-free water (RNasefreewater), the final volume is 20 μl , and incubated at 37°C for 1h.

[0085] 2) Add 1 μl of 0.5 μg / μl Oligo(dT) specific RT primer to the reaction tube, and incubate at 70° C. for 5 minutes.

[0086] 3) Immediately incubate on ice for at least 2 minutes to break the secondary structure of RNA and primers.

[0087] 4) Mix the ab...

Embodiment 3

[0103] Example 3 Expression of miRNA-1262 in acute myeloid leukemia cell lines

[0104] 1. Cell culture

[0105] Acute myeloid leukemia cell lines U937, THP, KG-1 and bone marrow mononuclear cells isolated from AML patients and healthy people were routinely cultured in RPMI1640 medium containing 10% fetal bovine serum at 37°C, 5% CO 2 in the incubator.

[0106] 2. QPCR

[0107] 2.1 Extraction of total cellular RNA: Extraction of total cellular RNA was performed using the RNA extraction kit from QIAGEN, following the instructions in the instructions.

[0108] 2.2 QPCR: the steps are the same as in Example 2.

[0109] 3. Results

[0110] Such as figure 2 As shown, compared with the bone marrow mononuclear cells of healthy people, the expression of miRNA-1262 in the bone marrow mononuclear cells of acute myeloid leukemia cell lines U937, THP, KG-1 and AML patients was significantly increased (P<0.05).

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Abstract

The invention discloses a miRNA marker which is miRNA-1262. Whether a patient suffers from acute myelogenous leukemia or not can be diagnosed by detecting the expression level of the miRNA-1262. As is proved by experiment, an acute myelogenous leukemia patient and a healthy person can be effectively distinguished through the miRNA-1262. On the basis, the miRNA-1262 can also be used for preparing acute myelogenous leukemia treatment drugs. The miRNA-1262 improves the sensitivity and specificity of acute myelogenous leukemia diagnosis greatly and provides a new drug target for gene therapy of acute myelogenous leukemia.

Description

technical field [0001] The invention belongs to the field of biomedicine, and relates to an acute myeloid leukemia miRNA marker and application thereof. Background technique [0002] Acute myeloid leukemia (AML) is a group of highly heterogeneous diseases, and its development is a multi-gene change and multi-step process. At present, cytogenetic abnormalities at the time of diagnosis have become a common clinical prognostic indicator for AML, but traditional There are still some defects in cytogenetics, and changes in molecular genetics are increasingly important for prognosis. In recent years, the incidence of AML has shown a gradual upward trend. Although the existing treatment methods can achieve a complete remission rate of 50% to 70%, the mortality rate of the disease is still high, and individualized stratified treatment for different patients has become the first priority. question. Comprehensive research on the role of genes on the occurrence and development of AML...

Claims

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Application Information

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IPC IPC(8): C12N15/113C12Q1/68A61K48/00A61P35/02
Inventor 边洋杨承刚
Owner QINGDAO MEDINTELL BIOMEDICAL CO LTD
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