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Leukemia-relevant circular circRNA-011235 gene and applications thereof

A leukaemia, ring technology, applied in the field of tumor molecular biology, can solve the problems of transplantation failure, hematopoiesis and its mechanism of action are unclear, and affect HSCs implantation and transplantation efficacy, etc.

Inactive Publication Date: 2016-12-07
THE THIRD XIANGYA HOSPITAL OF CENT SOUTH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] The injury of bone marrow mesenchymal stem cells (BMSCs) by TBI before bone marrow transplantation may affect the implantation and transplantation efficacy of HSCs, and even lead to transplantation failure. clear

Method used

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  • Leukemia-relevant circular circRNA-011235 gene and applications thereof
  • Leukemia-relevant circular circRNA-011235 gene and applications thereof
  • Leukemia-relevant circular circRNA-011235 gene and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1: Amplification of circular circRNA-011235 gene primer pair

[0020] According to the cDNA sequence shown in SEQ ID NO:1 of the circular circRNA-011235 gene, primers for amplification were designed, and the specific sequences of each primer are shown in Table 1.

[0021] Table 1 Circular circRNA-011235 Gene Amplification Primer Sequence

[0022] Primer name

[0023] After the above primers were artificially synthesized, they were used as a primer pair for amplifying the cDNA sequence of the circular circRNA-011235 gene.

Embodiment 2

[0024] Example 2: Preparation of total RNA in the sample

[0025] According to the RNA extraction steps of TRIZOL method, the total RNA in the bone marrow stromal cell samples of the control group and the whole body irradiation treatment group was extracted. A brief description is as follows:

[0026] 1. Homogenate

[0027] According to the instructions of the TRIZOL method, the sample was homogenized after adding TRIZOL reagent according to the number of cells;

[0028] 2. Two-phase separation

[0029] After incubating the homogenized sample at 15-30°C for 5 minutes, add 0.2ml of chloroform to every 1ml of TRIZOL reagent homogenized sample, and tightly cap the tube. Shake the tube vigorously by hand for 15 seconds, and incubate at 15 to 30°C for 2 to 3 minutes. Centrifuge at 12,000 x g for 15 min at 4°C. After centrifugation, the mixed liquid will be divided into the red phenol chloroform phase of the lower layer, the colorless aqueous phase of the middle layer and the u...

Embodiment 3

[0039] Embodiment 3: the synthesis of total cDNA sequence

[0040] The total RNA that passed the quality test was synthesized according to the method described below. The reagents used in the synthesis and their manufacturers are as follows:

[0041] RNase inhibitor (Epicentre); SuperScriptTM III Reverse Transcriptase (Invitrogen); 5×RT buffer (Invitrogen); 2.5mM dNTP mixture (dATP, dGTP, dCTP and dTTP each 2.5mM) (HyTest Ltd); Jun Biotechnology Co., Ltd.)

[0042] The thermocycler used for cDNA synthesis was Gene Amp PCR System 9700 (Applied Biosystems).

[0043] The specific operation method is as follows:

[0044] 1. Prepare the annealing mixture

[0045] RNA 800ng

[0046] 0.5ug / ul Random (N9) 1μl

[0047] dNTPs Mix (2.5mM) 1.6μl

[0048] Add RNase-free H 2 O to a total volume of 14.5 μl; the mixture was placed in a water bath at 65°C for 5 minutes, and placed on ice for 2 minutes.

[0049] 2. After short centrifugation, add RT reaction solution to the centrifuge ...

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Abstract

The invention discloses a leukemia-relevant circular circRNA-011235 gene and applications of the leukemia-relevant circular circRNA-011235 gene. According to the leukemia-relevant circular circRNA-011235 gene and the applications of the leukemia-relevant circular circRNA-011235 gene, the existence of the circRNA-011235 gene is proved first, through the detection on the expression condition of the gene in a patient under toal body irradiation, the result shows that the expression level of the circRNA-011235 gene is obviously increased. The marrow stroma cell transfected with the adenovirus vector with the overexpression of the circular circRNA-011235 gene is compared with a contrast marrow stroma cell transfected with a blank carrier, the result shows that the expression quantity of TGF-beta protein in the former cell is obviously increased, consequently, myelofibrosis is caused, then homing, self-renewal, proliferation and differentiation of hematopoietic cells are not facilitated, further, the curative effects of hematopoietic stem cell transplantation are influenced, therefore, the circular circRNA-011235 gene and the expression product of the circular circRNA-011235 gene can be taken as markers for predicating the hematopoietic stem cell transplantation curative effects, and can be taken as target genes and medicines for preparing leukemia medicines.

Description

technical field [0001] The invention belongs to the field of tumor molecular biology, and in particular relates to a circular circRNA-011235 gene related to leukemia and its application. Background technique [0002] Leukemia, commonly known as blood cancer, has posed a huge threat to people's health because of its unknown etiology, short survival period and high mortality rate. According to statistics, there are more than 4 million leukemia patients in my country, and more than 40,000 new cases will be added every year. Hematopoietic stem cell transplantation (HSCT) is currently the first choice to cure malignant hematological diseases, and total body irradiation (TBI) is one of the necessary auxiliary means for hematopoietic stem cell (HSCs) transplantation. The purpose of TBI treatment: one is immunosuppression, mainly to enable the transplanted bone marrow to be accepted by the recipient. The second is to eliminate malignant tumor cells in the body to achieve the purpo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12Q1/68
CPCC12N15/113C12N2310/10C12Q1/6886C12Q2600/118C12Q2600/136C12Q2600/158C12Q2600/178C12N2310/532
Inventor 聂新民张军华桂嵘黄蓉江静
Owner THE THIRD XIANGYA HOSPITAL OF CENT SOUTH UNIV
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