CN114214324A - miR-219a-2-3p and application thereof as molecular marker for early diagnosis of radiation damage - Google Patents
miR-219a-2-3p and application thereof as molecular marker for early diagnosis of radiation damage Download PDFInfo
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- CN114214324A CN114214324A CN202111394622.3A CN202111394622A CN114214324A CN 114214324 A CN114214324 A CN 114214324A CN 202111394622 A CN202111394622 A CN 202111394622A CN 114214324 A CN114214324 A CN 114214324A
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- C—CHEMISTRY; METALLURGY
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- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/11—DNA or RNA fragments; Modified forms thereof; Non-coding nucleic acids having a biological activity
- C12N15/113—Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing
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- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
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- C12N2310/00—Structure or type of the nucleic acid
- C12N2310/10—Type of nucleic acid
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- C12N2310/141—MicroRNAs, miRNAs
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- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/178—Oligonucleotides characterized by their use miRNA, siRNA or ncRNA
Abstract
The invention belongs to the technical field of biological detection, and relates to miR-219a-2-3p and application thereof as a molecular marker for early diagnosis of radiation damage. By using the miR-219a-2-3p as the molecular marker for early diagnosis of radiation damage, radiation exposure diagnosis can be conveniently, accurately and quantitatively carried out.
Description
Technical Field
The invention belongs to the technical field of biological detection, and relates to miR-219a-2-3p and application thereof as a molecular marker for early diagnosis of radiation damage.
Background
At present, the health monitoring of radiology workers mainly comprises health physical examination, health risk assessment, disease prevention and treatment and the like. The daily health examination mainly adopts routine clinical examination, and the special health examination comprises chromosome aberration and micronucleus analysis. However, the indexes related to the radiation-induced diseases cannot be effectively found in advance.
The molecular indexes obtained by the related technology of molecular biology can supplement and improve physical examination means, and early diagnosis and early treatment of diseases are realized. Therefore, in order to improve the existing health monitoring system and avoid or reduce the potential risk of radiation hazard, it is urgently needed to establish an early health monitoring technology for the radiology workers, so as to provide technical support and management basis for the post adaptability judgment of the radiology workers.
miRNA participates in regulation and control of cell proliferation and differentiation and apoptosis, and plays an important role in radiation-induced body injury. The physical properties of miRNA in human body fluid are very stable, and the miRNA is very suitable to be used as a biomarker.
miR-219a-2-3p is located on human chromosome 9, and the functions of the miR-219a-2-3p in tumor diagnosis and prognosis prediction are partially reported in the diagnosis and prognosis prediction of gastric cancer. However, the correlation research between the miR-219-2-3p and the ionizing radiation exposure is not reported at home and abroad.
Disclosure of Invention
The invention aims to provide miRNA, which can be used for conveniently, accurately and quantitatively performing radiation exposure diagnosis.
To achieve the purpose, in a basic embodiment, the invention provides a miRNA, wherein the miRNA is miR-219a-2-3p, and the sequence of the miRNA is shown as SEQ ID NO. 1.
The second purpose of the invention is to provide the application of miR-219a-2-3p as a molecular marker for early diagnosis of radiation damage, so that radiation exposure diagnosis can be conveniently, accurately and quantitatively carried out.
To achieve the purpose, in a basic embodiment, the invention provides the application of miR-219a-2-3p as a molecular marker for early diagnosis of radiation damage (namely the application of miR-219a-2-3p as the molecular marker in preparing a kit for early diagnosis of radiation damage).
In a preferred embodiment, the invention provides the use of miR-219a-2-3p as a molecular marker for early diagnosis of radiation injury, wherein the radiation injury is local or systemic tissue organ adverse reaction (such as nausea, vomiting, anorexia, leukopenia and/or skin redness and itching and the like).
The method has the beneficial effects that the miR-219a-2-3p disclosed by the invention is used as a molecular marker for early diagnosis of radiation damage, and radiation exposure diagnosis can be conveniently, accurately and quantitatively carried out.
Detailed Description
The following examples further illustrate the practice of the present invention, but the embodiments of the present invention are not limited to the following examples.
Example 1:
1. preliminary screening of radiation-sensitive molecular markers using high-throughput sequencing techniques
1) Extraction, quantification and quality inspection of cell total miRNA
Irradiating human peripheral venous blood with 2.0Gy gamma ray (simultaneously providing non-irradiated control group), extracting mRNA by using peripheral blood mRNA extraction kit (Tiangen Biochemical technology Co., Ltd.) 6h after irradiation, and performing quality detection and quantification on mRNA sample by using Nanodrop (OD 260/280: 1.81-1.87, total amount: 1699.50-3592.80 ng).
2) Construction of the library
Preparing a miRNA sequencing library by using the total RNA of each sample, and mainly comprising the following steps: 3' joint connection; 5' joint connection; synthesizing cDNA; PCR amplification; the 135-155bp PCR amplified fragment (corresponding to 15-35nt small RNA) was selected and subjected to library quality determination using Agilent 2100.
3) Sequencing
The pooled sequencing libraries of the different samples were denatured by 0.1M NaOH to generate single stranded DNA, captured on an ilmina flow cell, amplified in situ into clusters, and subjected to 51 cycles on an Illumina NextSeq 500 sequencer according to the supplier's instructions.
4) Data analysis
Differential expression analysis of miRNAs among groups is carried out by using edgeR, a threshold value is set to be 1.5-fold difference, a p value is less than or equal to 0.05, and miRNA with differential expression is screened, wherein a positive value of the difference fold indicates up-regulation, and a negative value indicates down-regulation.
According to analysis, compared with a control group, after 2.0Gy gamma ray irradiation, the expression of miR-219-2-3p is up-regulated by 2.511 times, and the p value is 0.009.
2. PCR verification confirms feasibility of miR-219-2-3p as molecular marker for early diagnosis of radiation injury
miRNA is extracted by a peripheral blood miRNA extraction kit (Tiangen Biochemical technology Co., Ltd.). And (3) carrying out reverse transcription on cDNA according to the instruction of a reverse transcription kit (Tiangen Biochemical technology Co., Ltd.), taking a proper amount of sample cDNA of a radiation irradiation group and a control group, and carrying out PCR verification on miR-219-2-3p differential expression. The primer sequences and PCR amplification conditions are shown in Table 1. Further PCR verification of the DNA sequence shows that: the validation results were consistent with the sequencing results, see table 2.
TABLE 1 primer sequences and amplification reaction conditions for RT-PCR amplification of miRNA
TABLE 2 relative quantification of RT-PCR amplification
The experimental results show that the expression level of miR-219-2-3p in human peripheral blood is obviously increased compared with a control group after the action of ionizing radiation. Therefore, miR-219-2-3p is screened as a molecular marker for diagnosing early damage caused by ionizing radiation exposure and is used as an index for health monitoring of the irradiated personnel.
It will be apparent to those skilled in the art that various changes and modifications may be made in the present invention without departing from the spirit and scope of the invention. Thus, if such modifications and variations of the present invention fall within the scope of the claims of the present invention and their equivalents, the present invention is intended to include such modifications and variations. The foregoing examples or embodiments are merely illustrative of the present invention, which may be embodied in other specific forms or in other specific forms without departing from the spirit or essential characteristics thereof. The described embodiments are, therefore, to be considered in all respects as illustrative and not restrictive. The scope of the invention should be indicated by the appended claims, and any changes that are equivalent to the intent and scope of the claims should be construed to be included therein.
Sequence listing
<110> China institute for radiation protection
<120> miR-219a-2-3p and application thereof as molecular marker for early diagnosis of radiation damage
<130> 21F0705CN
<141> 2021-11-23
<160> 1
<170> SIPOSequenceListing 1.0
<210> 1
<211> 22
<212> RNA
<213> human (Homo sapiens)
<400> 1
agaauugugg cuggacaucu gu 22
Claims (3)
1. A miRNA, comprising: the miRNA is miR-219a-2-3p, and the sequence of the miRNA is shown in SEQ ID NO. 1.
Use of miR-219a-2-3p as a molecular marker for early diagnosis of radiation injury.
3. Use according to claim 2, characterized in that: the radiation injury is local or systemic tissue organ adverse reaction.
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Citations (6)
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| CN103805696A (en) * | 2013-12-12 | 2014-05-21 | 焦志军 | Micro RNA (Ribonucleic Acid) molecular marker for diagnosing rheumatoid arthritis and detection kit thereof |
| CN110261518A (en) * | 2019-06-24 | 2019-09-20 | 南华大学 | A kind of screening of human body effect of low dose radiation molecular injury marker taurine and verification method |
| CN111172272A (en) * | 2020-01-17 | 2020-05-19 | 中国辐射防护研究院 | Application of TRIP12 gene as molecular marker for judging susceptibility to radiation damage |
| US20200239967A1 (en) * | 2017-10-03 | 2020-07-30 | Geneseq Pty. Ltd. | Method of diagnosis, staging and monitoring of melanoma using microrna gene expression |
| CN111826443A (en) * | 2020-07-03 | 2020-10-27 | 清华大学深圳国际研究生院 | Application of serum exosome micro RNAs and liver cancer detection kit |
| CN112126686A (en) * | 2020-10-23 | 2020-12-25 | 河北仁博科技有限公司 | Early screening molecular marker for renal fibrosis and application thereof |
-
2021
- 2021-11-23 CN CN202111394622.3A patent/CN114214324A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103805696A (en) * | 2013-12-12 | 2014-05-21 | 焦志军 | Micro RNA (Ribonucleic Acid) molecular marker for diagnosing rheumatoid arthritis and detection kit thereof |
| US20200239967A1 (en) * | 2017-10-03 | 2020-07-30 | Geneseq Pty. Ltd. | Method of diagnosis, staging and monitoring of melanoma using microrna gene expression |
| CN110261518A (en) * | 2019-06-24 | 2019-09-20 | 南华大学 | A kind of screening of human body effect of low dose radiation molecular injury marker taurine and verification method |
| CN111172272A (en) * | 2020-01-17 | 2020-05-19 | 中国辐射防护研究院 | Application of TRIP12 gene as molecular marker for judging susceptibility to radiation damage |
| CN111826443A (en) * | 2020-07-03 | 2020-10-27 | 清华大学深圳国际研究生院 | Application of serum exosome micro RNAs and liver cancer detection kit |
| CN112126686A (en) * | 2020-10-23 | 2020-12-25 | 河北仁博科技有限公司 | Early screening molecular marker for renal fibrosis and application thereof |
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