Application and detection method of circRNA0003307 gene

A gene and gene expression technology, applied in the direction of DNA / RNA fragments, recombinant DNA technology, microbial measurement / inspection, etc., can solve the problem of less circRNA in AS patients, and achieve enrichment of pathogenesis, phosphorylation and nuclear translocation reduction , improve the effect of the pathogenesis

Active Publication Date: 2021-01-12
FIRST AFFILIATED HOSPITAL OF ANHUI UNIV OF CHINESE MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, there are few studies on circRNA in AS patients

Method used

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  • Application and detection method of circRNA0003307 gene
  • Application and detection method of circRNA0003307 gene
  • Application and detection method of circRNA0003307 gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0020] In order to explore the expression of circRNA0003307 and the expression of regulatory proteins (PI3K, AKT, TNFAIP2 and IL-1β) in peripheral blood mononuclear cells of patients with AS.

[0021] After the project passed the review of the Ethics Committee, the applicant systematically studied the new diagnostic and therapeutic targets of human AS, extracted the peripheral blood of AS patients, extracted PBMCs, and detected the expression of circRNA0003307 gene by real-time PCR.

[0022] In this embodiment, a total of 30 blood samples from patients with active AS and 30 blood samples from healthy people in the control group were collected, a total of 60 samples (see Table 1 for specific information from 30 patients, and Table 2 for sample characteristics).

[0023] Table 1 Sample information

[0024]

[0025]

[0026] Table 2 Sample Characteristics

[0027]

[0028] In this example, 30 cases of AS and 30 cases of healthy control mononuclear cells derived from per...

Embodiment 2c

[0041] Example 2 circRNA0003307 interference sequence and its application

[0042] This example designs and synthesizes its specific interference sequence si-circRNA0003307 (its nucleotide sequence is shown in SEQ ID NO.3) for the full-length sequence of circRNA0003307, and infects synovial fibroblasts to inhibit the circRNA0003307 gene in the cells expression, and further apply the interference gene carrier to regulate the inflammatory response of synovial fibroblasts.

[0043] The specific cell infection method is as follows:

[0044] 1. siRNA transfection steps

[0045] 1) Seed plate, the cell density is 50% suitable.

[0046] 2) The next day (24-36 hours later), the transfection method of each well is as follows:

[0047] A Dissolve 20pmol siRNA in 50ul Opti-mem serum-free medium; B dissolve 1ul lipo2000 in 50ul Opti-mem serum-free medium, mix well and place at room temperature for 5min; C mix tubes A and B and place for 20min.

[0048] 3) During the transfection perio...

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Abstract

The invention discloses application and a detection method of a circRNA0003307 gene, and belongs to the technical field of molecular biology of rheumatism. A preparation for detecting the expression quantity of the circRNA0003307 gene comprises a primer pair with nucleotide sequences as shown in SEQ ID NO. 1 and NO. 2, and the preparation for detecting the expression quantity of the gene can be applied to preparation of an ankylosing spondylitis detection preparation. A specific interference sequence si-circRNA0003307can significantly inhibit the expression level of circRNA0003307 in synovialcells, and can cause that the proliferation ability of the synovial cells is obviously weakened compared with the previous proliferation ability, LPS-induced inflammatory cytokines are reduced, and PI3K and AKT phosphorylation and nuclear translocation are obviously reduced. The discovery is expected to further enrich and perfect the research of AS pathogenesis, and also brings hope for developingnovel biomarkers for AS early diagnosis, targeted therapy and prognosis evaluation.

Description

technical field [0001] The invention belongs to the technical field of molecular biology of rheumatic diseases, and relates to the expression, application and detection field of a key circular RNA gene (circRNA0003307), specifically an application of the circRNA0003307 gene and a detection method thereof. Background technique [0002] Ankylosing spondylitis (AS) is a common inflammatory rheumatic disease affecting the axial skeleton, causing characteristic inflammatory back pain that can lead to structural and functional impairment and reduced quality of life (Jürgen Braun, Sieper J. Ankylosing Spondylitis[J].The Lancet,2007,369(9570):1379-1390.), often accompanied by eyeball uveitis, heart valve disease, lung, intestinal and other systemic diseases (Smith JA.Update on Ankylosing Spondylitis: Current Concepts in Pathogenesis. Current Allergy & Asthma Reports. 2015; 15(1): 489; Ciccia F, Guggino G, Rizzo A, et al. Dysbiosis and zonulin upregulation alter gut epithelial and va...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12N15/113
CPCC12Q1/6883C12Q2600/178C12Q2600/158
Inventor 刘健姜辉万磊龙琰黄旦
Owner FIRST AFFILIATED HOSPITAL OF ANHUI UNIV OF CHINESE MEDICINE
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