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Application of SNP bite of CLIP3 gene

A site, aortic dissection technology, applied in the field of biomedical detection, can solve the problems of high AD vascular risk, difficult early risk assessment, and difficult early detection

Active Publication Date: 2018-12-28
中国医学科学院阜外医院深圳医院
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] AD has a high vascular risk and is not easy to detect early. The positive rate of genetic diagnosis based on known disease-causing genes is limited, making early risk assessment difficult, and it is urgent to discover new molecular markers

Method used

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  • Application of SNP bite of CLIP3 gene
  • Application of SNP bite of CLIP3 gene
  • Application of SNP bite of CLIP3 gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1 Sample collection

[0030] From January 2017 to April 2018, 59 patients with Stanford type A aortic dissection were diagnosed by aortic CTA in Shenzhen Sun Yat-sen Cardiovascular Hospital, and 2 mL of whole blood and 590 normal control samples were collected from the searched by Makino Sequencing Company. database. Informed consent was obtained from the patient and reviewed by the Ethics Committee.

[0031] Sample processing: EDTA anticoagulated whole blood was mixed with Trizol at a ratio of 1:1, mixed thoroughly and placed in a 1.8mL cell cryopreservation tube, cooled rapidly in liquid nitrogen for 30 seconds and stored in a -80°C refrigerator.

Embodiment 2

[0032] Embodiment 2 extracts blood sample DNA

[0033] (1) Add 1 mL of CL cell lysate to 1 mL of EDTA (0.01M, China, Huamei Bioengineer) anticoagulated blood, gently invert and mix 6 times, centrifuge at 3600 rpm for 5 min, and discard the supernatant;

[0034] (2) Pour 1mL of CL cell lysate into the centrifuge tube again, gently invert and mix 6 times, centrifuge at 3600rpm for 5min, discard the supernatant; under the premise of ensuring that the precipitate remains in the tube, invert the centrifuge tube Stand on clean absorbent paper for 2 minutes;

[0035] (3) configuring a mixed solution of proteinase K and buffer FG;

[0036] (4) Add 500 μL of proteinase K and buffer FG mixture, then mix until the solution has no lumps;

[0037] (5) Water bath at 65°C for 30 minutes, and mix by inverting several times during the period;

[0038] (6) Add 1 mL of isopropanol, then invert and mix until clustered or filamentous genomic DNA appears;

[0039] (7) Centrifuge at a speed of 3...

Embodiment 3

[0047] Example 3 Whole Exome Sequencing

[0048] 1. DNA library construction

[0049] 3 μg of DNA was ultrasonically fragmented, the ends of the interrupted fragments were blunted, A was added to the 3' end, and the adapter was connected, and the fragments between 350 and 400 bp were selected to prepare the genome-wide library. Library samples were quality controlled using an Agilent 2100 bioanalyzer (Agilent Technologies, USA).

[0050] 2. Target region capture sequencing

[0051] Whole-exome detection was performed using GenCap liquid-phase capture target gene technology (Beijing Mikino Company). Mix 1 μg of DNA library with BL buffer and probe, heat at 95°C for 7 minutes, heat at 65°C for 2 minutes, add 23 μL of HY buffer preheated to 65°C, and hybridize at 65°C for 22 hours. 50 μL MyOne magnetic beads (Life Technology, USA) were washed 3 times with 500 μL 1X binding buffer, and resuspended in 80 μL 1X binding buffer. Add 64 μL 2X Binding Buffer to the hybridization mix...

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Abstract

The invention discloses a full-exon sequence detecting technology screening based and aortic dissection disease related SNP bite. The SNP bite comprises the following aortic dissection easily-sensed SNP bites: CLIP3: NM_001199570: exon1: c. 32C) T: p. P11L. The invention further provides application of the SNP bite in a detection product for detecting aortic dissection disease. According to the application, the aortic dissection easily-sensed SNP bite of SEQ ID NO.3 is found out, and the application prospect thereof in early prediction of the aortic dissection is researched. The gene bite related to the aortic dissection disease is possibly an early biomarker of the aortic dissection; and a new direction is supplied to further research the genetic molecular mechanism of the aortic dissection and explore a medicine target for early prevention and treatment of the aortic dissection.

Description

technical field [0001] The invention relates to the technical field of biomedical detection, in particular to the application of the SNP site of the CLIP3 gene related to aortic dissection disease. Background technique [0002] Aortic dissection (AD) refers to a series of external factors (such as high blood pressure, trauma, etc.), based on the presence or absence of lesions in the aortic wall itself, a breach in the aortic intima occurs. Blood in the aortic intimal breach invades the middle layer of the aortic wall, and the aortic middle layer is continuously torn and separated longitudinally, resulting in a state of coexistence of true and false aortic lumens. Aortic dissection has sudden onset, rapid development, and severe prognosis. Despite the continuous improvement of various current treatment methods, the morbidity and mortality of AD are still high, and if proper and timely treatment is not carried out, the mortality rate of AD is extremely high. It has been repo...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883
CPCC12Q1/6883C12Q2600/118C12Q2600/156
Inventor 蒲佐
Owner 中国医学科学院阜外医院深圳医院
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