Human intron derived 27 base microRNA and application thereof to blood pressure regulation

An intron and base technology, applied in the field of rat tail vein injection and gene recombination, can solve the problems of unknown etiology of hypertension, unclear mechanism of pathological development, lack of disease prevention and treatment measures, etc.

Pending Publication Date: 2020-02-28
GUANGXI INT ZHUANG MEDICINE HOSPITAL
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, the etiology of hypertension is unknown, and the mechanism of its pathological development is also unclear, so there is a lack of prevention and treatment measures to completely and efficiently cure this disease

Method used

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  • Human intron derived 27 base microRNA and application thereof to blood pressure regulation
  • Human intron derived 27 base microRNA and application thereof to blood pressure regulation
  • Human intron derived 27 base microRNA and application thereof to blood pressure regulation

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0031] Example 1 Construction and detection of human intron-derived 27-base microRNA high-expression plasmid

[0032] The human intron-derived 27-base microRNA nucleotide sequence was obtained through the Blast system in NCBI, and its nucleotide sequence is shown in SEQ ID NO.1; the target gene was amplified by PCR, and the target gene was obtained under the PCR reaction system Gene fragment. After double digestion and linear ligation, use a small amount of plasmid extraction kit (Beijing Tiangen Biochemical Technology Co., Ltd.) to extract the plasmid, and take the positive clone liquid and send it to the company for sequencing identification. After the verification is correct, a large amount of amplification and extraction is carried out to obtain a human intron-derived 27-base microRNA high-expression plasmid (see figure 1 ).

[0033] Upstream amplification primer: 5'-GGAAGTCTAGACCTGCTGCA-3',

[0034] Downstream amplification primer: 5'-GTCGGTGTCGTGGAGTCG-3';

[0035] A...

Embodiment 2

[0038] Example 2 Transfection and transfection determination of human intron-derived 27-base microRNA high-expression plasmid in rat aortic tissue

[0039] The animals were randomly divided into two groups, 10 in each group, namely the 27-base microRNA group and the blank plasmid group. Rats in the 27-base microRNA group were mixed with 27-nt-miRNA high-expression plasmid and liposome at a mass ratio of 3:1, diluted with saline, and injected into the body through the tail vein, and rats in the blank plasmid group were treated with blank plasmid Inject into the body through the tail vein. The dosage of both groups was 100ng plasmid DNA / kg body weight, injected once every two days, and injected continuously for 15 weeks. Rats were kept in a suitable environment and allowed to eat and drink normally during this period.

[0040] After 15 weeks, the animals were sacrificed, and the aortic tissues of the rats were taken, frozen at -80°C, frozen sections were made, and the green flu...

Embodiment 3

[0041] The mensuration of embodiment 3 rat blood pressure

[0042] Before and after the injection, the systolic blood pressure (SBP), diastolic blood pressure (DBP), mean blood pressure (MBP) and heart rate (HR) of each rat were measured using a rat tail artery non-invasive blood pressure measuring instrument. Measure blood pressure at least 5 times in a row and take the average value. Measured twice a week, continuous observation for 15 weeks. During the experiment, the rats were given normal diet, water, and a suitable environment. The results showed that the SBP, DBP and MBP of animals in the 27-base microRNA group were all increased. The SBP, DBP and MBP of animals in the blank plasmid group were slightly elevated at first, and then gradually decreased to normal levels after the 10th week (such as figure 2 ); It shows that human intron-derived 27-base microRNA can increase blood pressure in normal rats.

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Abstract

The invention discloses a human intron derived 27 base microRNA and an application thereof to blood pressure regulation. The human intron derived 27 base microRNA is generated by four-time duplicationof a fourth intron 27 base duplicon of eNOS (endothelial nitric oxide synthase), and the nucleotide sequence of the human intron derived 27 base microRNA is as shown in SEQ ID NO. 1. A high-expression plasmid is prepared according to the sequence, the high-expression plasmid and lipidosome are mixed according to the weight ratio of 3:1 and diluted by normal saline, and mixture is injected into the body of an SD (sprague-daw) rat through rat caudal veins to prove that the human intron derived 27 base microRNA has a regulating effect on rat blood pressure, can be used for researching hypertension diseases, finally can be used for screening some antihypertensive drugs and preparing a hypertension early warning chip and has a potential application prospect.

Description

technical field [0001] The invention belongs to the technical field of rat tail vein injection and gene recombination, and specifically relates to a human intron-derived 27-base microRNA and its application in regulating blood pressure. Background technique [0002] Hypertension is the most common cardiovascular disease, often causing serious heart, brain, and kidney complications, affecting 20% ​​to 50% of adults worldwide. According to statistics, there are about 200 million hypertensive patients in my country at present, and 1 out of every 5 adults suffers from high blood pressure, accounting for about 1 / 5 of the total number of people with high blood pressure in the world. Nearly 7.6 million patients worldwide die of this disease every year . The World Health Organization (WHO) defines hypertension as systolic blood pressure ≥ 140 mmHg and / or diastolic blood pressure ≥ 90 mmHg in the absence of antihypertensive drugs. Hypertension is divided into 1, 2, and 3 grades: 1) ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/85C12Q1/6883A61K31/7105A61P9/12
CPCC12N15/113C12N15/85C12Q1/6883A61P9/12C12N2310/141C12N2800/107C12Q2600/158C12Q2600/178
Inventor 欧和生杨鹏罗雪兰
Owner GUANGXI INT ZHUANG MEDICINE HOSPITAL
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