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Micro ribonucleic acid (miRNA) marker group related to acute myelogenous leukemia, and specific primer and application of marker group

An acute myeloid, specific primer technology, applied in the field of medical molecular biology, can solve problems such as non-involvement, achieve the effects of strong complementarity, accurate detection and quantification, and improved sensitivity and specificity

Active Publication Date: 2015-06-10
SHANDONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the existing early diagnostic markers for AML do not involve the application of the combination of miR-23b-3p and miR-26a-5p based on changes in granulocyte miRNA expression profiles as early diagnostic molecular markers for AML.

Method used

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  • Micro ribonucleic acid (miRNA) marker group related to acute myelogenous leukemia, and specific primer and application of marker group
  • Micro ribonucleic acid (miRNA) marker group related to acute myelogenous leukemia, and specific primer and application of marker group
  • Micro ribonucleic acid (miRNA) marker group related to acute myelogenous leukemia, and specific primer and application of marker group

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] 1. 12 cases of acute myeloid leukemia and 20 cases of normal healthy controls received physical examination, and 5ml of peripheral blood was extracted using an anticoagulant tube to separate and obtain granulocytes.

[0031] 2. Extraction of total RNA from granulocytes: TRI Reagent BD reagent was used to extract RNA from granulocytes.

[0032] (1) Lysis: 0.75ml TRI Reagent BD to prepare the lysate;

[0033](2) Two-phase separation: lysate + 0.1ml bromochloropropane or 0.2ml chloroform;

[0034] (3) RNA precipitation: aqueous phase layer + 0.5ml isopropanol;

[0035] (4) Wash RNA: 1ml 75% ethanol;

[0036] (5) RNA dissolution: Dissolve RNA with FORMAzol, water or a 0.5wt% SDS solution prepared by blowing and mixing with a pipette tip.

[0037] 3. miRNA chip hybridization: using Exiqon miRCURY LNA TM The 11.0 chip includes all miRNAs from the three species of human, mouse, and rat in the miBase database. The capture probes on the chip are all patented LNA probes, whic...

Embodiment 2

[0049] Using the specific primers of miR-26a-5p and miR-23b-3p (as shown in Table 3), miR-26a-5p and miR-26a-5p and There are significant differences in the expression of miR-23b-3p between the acute myeloid leukemia case group and the healthy control group, as shown in image 3 shown.

[0050] Table 3. Primer information for related miRNAs (Note: GSP is the corresponding specific primer, R is the primer that matches the RT primer)

[0051]

[0052] According to the above results, by analyzing the expression levels of miRNAs in two groups of peripheral blood granulocyte samples (case group and healthy control group), 90 percent of the expression levels of miR-23b-3p and miR-26a-5p in the healthy control group were respectively The median is the threshold value, using miR-23b-3p and miR-26a-5p to group, the expression level is less than the 90th percentile is scored as 0 points, and it is divided into the healthy group, and the expression level is greater than or equal to t...

Embodiment 3

[0060] Auxiliary early diagnosis kit for acute myeloid leukemia, including:

[0061] The nucleotide sequence is as the specific primer of the miR-23b-3p marker shown in SEQ ID NO.3 and SEQ ID NO.4; the nucleotide sequence is as the miR shown in SEQ ID NO.5 and SEQ ID NO.6 -Specific primers for the 26a-5p marker;

[0062] Reverse transcriptase (purchased from Takara), buffer (purchased from Takara), dNTPs (purchased from Takara), MgCl 2 (purchased from Shanghai Sangong), DEPC water and Taq enzyme (purchased from Takara Company), and human U6miRNA.

[0063] The process of using the above kit is as follows: (1) miRNA differential expression profile analysis: select acute myeloid leukemia cases (N>20) and healthy controls (N>20) matched with their gender and age, and detect their peripheral blood granulocyte miRNA Expression profile and content, analyze the commonality and characteristics of miRNAs between cases and healthy controls, and screen specifically expressed miRNAs. (2...

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Abstract

The invention relates to a micro ribonucleic acid (miRNA) marker group related to acute myelogenous leukemia, and a specific primer and application of the miRNA marker group. The marker group is composed of a miR-23b-3p marker and a miR-26a-5p marker, wherein a nucleotide sequence of the miR-23b-3p marker is as shown in SEQ ID NO.1, and the nucleotide sequence of the miR-26a-5p marker is as shown in SEQ ID NO.2. The invention also relates to a specific primer and application of the miRNA marker group. Two markers screened by the micro ribonucleic acids (microRNAs) marker group provided by the invention are strong in complementarity. Thus, the detection is quantitative and accurate, and the sensibility and the specificity of disease diagnosis are greatly improved.

Description

technical field [0001] The invention relates to an acute myeloid leukemia-related miRNA marker group, specific primers and applications thereof, and belongs to the technical field of medical molecular biology. Background technique [0002] Acute myeloid leukemia (AML) is a heterogeneous myeloid tumor, also known as acute non-lymphocytic leukemia, which often progresses rapidly. Its characteristic is that the normal differentiation of hematopoietic stem cells is blocked, and the clonal proliferation of malignant tumor cells replaces normal hematopoiesis, resulting in a series of symptoms such as anemia, hemorrhage, infection and organ infiltration. Among the mortality rates caused by malignant tumors, leukemia ranks first among children and adults under the age of 35. In my country, among leukemias, acute myeloid leukemia has the highest annual incidence rate, accounting for about 58.7% of all cases. [0003] Acute myeloid leukemia not only requires early and timely diagnos...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6886C12Q2600/158C12Q2600/178
Inventor 李曦
Owner SHANDONG UNIV
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