Diagnosis molecular marker circRNA, kit and application of idiopathic optic neuritis

A technology of molecular markers and optic neuritis, applied in the direction of DNA/RNA fragments, recombinant DNA technology, biochemical equipment and methods, etc., can solve the problems of few circRNAs and unreported research, and achieve good stability, not easy to degrade, The effect of high auxiliary diagnostic value

Active Publication Date: 2021-07-02
THE SECOND XIANGYA HOSPITAL OF CENT SOUTH UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are still few studies on circRNAs related to idiopathic optic neuritis, and no studies using circRNAs as biomarkers have been reported.

Method used

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  • Diagnosis molecular marker circRNA, kit and application of idiopathic optic neuritis
  • Diagnosis molecular marker circRNA, kit and application of idiopathic optic neuritis
  • Diagnosis molecular marker circRNA, kit and application of idiopathic optic neuritis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0040] Example 1 Establishment of circRNA expression profile in idiopathic optic neuritis

[0041] Sample Collection:

[0042] This study has been approved by the Ethics Committee of the Second Xiangya Hospital of Central South University. All cerebrospinal fluid samples were obtained by lumbar puncture: instruct the patient to take the thoraco-knee position, take the L3 / 4 intervertebral space as the puncture point, routinely disinfect, puncture after layer-by-layer local anesthesia with 2% lidocaine, use a common tube to collect 1ml of cerebrospinal fluid, and freeze it The tubes were aliquoted in 250ul per tube and stored in a -80°C refrigerator within 15 minutes of isolation for circRNA biomarker analysis. Inclusion criteria for the control group: 1. Clinical diagnosis of headache and cause 2. No abnormalities in the head MRI examination. 3. No abnormalities in the biochemical, routine, and three major stainings of the cerebrospinal fluid examination. 4. No ophthalmic dise...

Embodiment 2

[0051] Embodiment 2 qRT-PCR verification

[0052] In the previously established expression profiles, 14 target circRNAs with significant differential expression were selected and verified again in 10 samples by qPT-PCR. The differential expression of the five circRNAs in the cerebrospinal fluid of the two groups was statistically different, and the change trend was consistent with that of the microarray, respectively, hsa_circRNA_0007503 and hsa_circRNA_007630 were up-regulated compared with the control group; hsa_circRNA_406587, hsa_circRNA_054220 and hsa_circRNA_005133 were down-regulated compared with the control group ( image 3 ). Then select the circRNA with statistical difference: hsa_circRNA_0007503 for further verification.

Embodiment 3

[0053] Example 3 Using qRT-PCR to further expand sample size verification

[0054] Expression level of hsa_circRNA_0007503 in cerebrospinal fluid to test its diagnostic value

[0055] Sample collection and total RNA extraction were the same as before.

[0056] cDNA synthesis and qRT-PCR:

[0057] Use RevertAidFirst Strand cDNA Synthesis Kit (Thermofisher) to prepare the reverse transcription system according to the manual for reverse transcription. Reverse transcription products can be stored at -20°C or used immediately. Download the circRNA sequence through the circbase database or UCSC genome browser, and use the primer design software Primer 5.0 to design primers for the target sequence and internal reference sequence. (Table 1) qPCR detection was performed using 2X FastStart Universal SYBR Green Master (Roche). The 96-well plate was placed in a Realtime PCR instrument (StepOnePlusTM Real-Time PCR System, Applied Biosystems) on the reaction. The conditions were 95° C...

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Abstract

The invention relates to the field of biological detection, in particular to circRNA, a kit and application of a diagnostic molecular marker for idiopathic optic neuritis. The present invention adopts circRNA chip detection to obtain abnormally expressed cerebrospinal fluid circRNAs expression profile, and uses qRT-PCR method for verification, and has strict design and mature evaluation system. It is the first time to discover hsa_circ_0007503 in cerebrospinal fluid as an important biological detection index. The detection index is minimally invasive and cheap, and can be widely used in clinical detection to solve the problems of long clinical diagnosis time, cumbersome process, high cost and low sensitivity of idiopathic optic neuritis . It provides new ideas for the diagnosis of idiopathic optic neuritis, and at the same time provides a new direction for improving the pathogenesis and therapeutic targets of idiopathic optic neuritis.

Description

technical field [0001] The invention relates to the field of biological detection, in particular to circRNA, a kit and application of a diagnostic molecular marker for idiopathic optic neuritis. Background technique [0002] Optic neuritis refers to acute optic nerve inflammatory lesions caused by various etiologies, and is the most common blinding optic nerve disease among young and middle-aged people. It is mainly manifested by acute or subacute vision loss and eye movement pain. The classification includes idiopathic, infection-related, autoimmune-related and other types. Among them, idiopathic optic neuritis is the most common type. Its etiology is complicated, some patients relapse, and the prognosis is relatively poor. It is one of the important reasons for loss, which has a serious impact on the physical and mental health of patients. Clinically, in order to accurately diagnose idiopathic optic neuritis, it is necessary to differentiate and exclude various diseases,...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/6883C12Q1/686C12N15/113
CPCC12Q1/686C12Q1/6883C12Q2600/158C12Q2600/178C12Q2561/113C12Q2563/107C12Q2545/114
Inventor 江冰陈碧玥卢伟王聪罗莹莹张璐丝刘利群周也荻张楠邹京伶段向巍
Owner THE SECOND XIANGYA HOSPITAL OF CENT SOUTH UNIV
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