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Application of ARHGAP26 gene to preparation of Parkinson diagnostic tool

A Parkinson's disease and tool technology, applied in the field of molecular diagnosis, can solve the problems of slow progress in pathological mechanism research, achieve timely gene diagnosis and reduce mortality

Inactive Publication Date: 2018-05-15
QINGDAO MEDINTELL BIOMEDICAL CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because the pathological changes of neurodegenerative diseases occur in the brain, the brain tissue is obtained after autopsy, so the research on the pathological mechanism is slow

Method used

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  • Application of ARHGAP26 gene to preparation of Parkinson diagnostic tool
  • Application of ARHGAP26 gene to preparation of Parkinson diagnostic tool

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Screening for genes differentially expressed in Parkinson's disease patients and normal individuals

[0037] 1. Research object:

[0038] Collect 10 cases of primary PD patients, 5 males and 5 females, aged 32-85 years, with a course of disease ranging from 5 months to 20 years. PD inclusion criteria: all diagnostic criteria meet the clinical diagnostic criteria for PD (refer to "Jiang Yuping, Wang Jian, Ding Zhengtong, et al., Diagnostic criteria for primary Parkinson's disease, 2005, Chinese Clinical Neuroscience, 2006, 14:40" ). Exclusion criteria: (1) essential tremor; (2) secondary parkinsonism; (3) severe dementia, dysarthria; (4) patients with other mental diseases. This study was approved by the Hospital Ethics Committee and all patients signed informed consent.

[0039] Normal group: 10 healthy volunteers aged 32-80 were selected, 5 males and 5 males.

[0040] There was no significant difference in age and gender between the two groups (P>0.10), w...

Embodiment 2

[0066] Embodiment 2QPCR experiment verifies the genes differentially expressed in patients with Parkinson's disease and normal people

[0067] 1. Research object:

[0068] The screening criteria were the same as in Example 1, 50 cases of Parkinson's disease patients and 50 normal persons.

[0069] 2. Extraction of total RNA in blood

[0070] Step is with embodiment 1.

[0071] 3. Reverse transcription

[0072] 1 μg of total RNA was reverse-transcribed to synthesize cDNA using reverse transcription buffer. Use 25μl reaction system, take 1μg total RNA for each sample as template RNA, add the following components to the PCR tube respectively: DEPC water, 5× reverse transcription buffer, 10mmol / L dNTP, 0.1mmol / l DTT, 30μmmol / l Oligo dT, 200 U / μl M-MLV, template RNA. Incubate at 42°C for 1h, then centrifuge briefly at 72°C for 10min.

[0073] 4. QPCR

[0074] (1) Primer design

[0075]QPCR amplification primers were designed according to the coding sequences of ARHGAP26 gen...

Embodiment 3

[0091] Example 3 Verification of expression products of differentially expressed genes in patients with Parkinson's disease and normal people by immunoblotting experiments

[0092] 1. Clinical object: same as embodiment 2.

[0093] 2. Mononuclear cell isolation

[0094] Take 10ml of venous blood from patients with Parkinson's disease and normal people, inject it into a sterile vial containing heparin, and shake it gently immediately after capping. Add an equal volume of HBSS (NaCl 8.0g, NaCl 2 HPO 4 0.132g, KH 2 PO 4 0.06g, KCl0.4g, phenol red 1ml, NaHCO 3 0.35g, D-glucose 1.0g, dissolved in 1000ml double distilled water), to reduce the aggregation of red blood cells. Draw 8ml of lymphocyte stratification solution into a 50ml centrifuge tube, slowly add the diluted blood along the tube wall, keep the interface clear, do not mix the two, centrifuge at 20°C 2000r / min for 30min, carefully absorb the stratification solution and plasma transfer The turbid off-white layer,...

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Abstract

The invention discloses an ARHGAP26 gene capable of being taken as a molecular marker for early diagnosis of a Parkinson's disease. The invention discovers that significant differences exist in the expressions of the ARHGAP26 gene in blood of an average person and a patient with the Parkinson's disease by utilizing a gene chip and a QPCR method, that is, whether a subject suffers from the Parkinson's disease or not can be judged by detecting the expression situation of the ARHGAP26 gene in the blood. According to the correlation between the both, the invention develops a kit for diagnosing theParkinson's disease, the kit is used for the diagnosis of the Parkinson's disease by detecting the expression of the ARHGAP26 gene. The diagnostic kit developed by the invention can be used for earlydiagnosis on diseases and has a wide application prospect clinically.

Description

technical field [0001] The invention belongs to the field of molecular diagnosis, and relates to a molecular marker for the diagnosis of Parkinson's disease, in particular to the application of the molecular marker-ARHGAP26 gene in blood in the preparation of products for diagnosing Parkinson's disease. Background technique [0002] PD is the second most common, slowly occurring, age-related neurodegenerative disease after Alzheimer's disease, also known as tremor paralysis. Pathologically, the selective loss of dopaminergic neurons (DNs) in the substantia nigra of the midbrain, the formation of Lewy body (LB) in the remaining neurons, and the significant reduction of dopamine content in the striatum lead to resting tremor, Myotonia, bradykinesia, and abnormal posture and gait are the four main clinical symptoms (Braak H, Tredici K D, et al. Staging of brain pathology related to sporadic Parkinson, sdisease. Neurobiology of Aging, 2003, 24 (2): 197 -211). In industrialized...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883G01N33/68
CPCC12Q1/6883C12Q2600/158G01N33/6896G01N2333/47G01N2800/2835G01N2800/50
Inventor 汪冰怡肖枫
Owner QINGDAO MEDINTELL BIOMEDICAL CO LTD
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