Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Adeno-associated virus recombinant vector for knocking out CXCL12 gene and construction method and use thereof

A technology of recombining vectors and viruses, applied in the direction of viruses/phages, using vectors to introduce foreign genetic material, viruses, etc., can solve the problems of lack of effective treatment methods and methods for glioma genes

Active Publication Date: 2019-01-18
汉恒生物科技(上海)有限公司
View PDF2 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The problem in the prior art is the lack of genetic means and methods for effective treatment of glioma

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Adeno-associated virus recombinant vector for knocking out CXCL12 gene and construction method and use thereof
  • Adeno-associated virus recombinant vector for knocking out CXCL12 gene and construction method and use thereof
  • Adeno-associated virus recombinant vector for knocking out CXCL12 gene and construction method and use thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] 1. pHBAAV-CMV-Sacas9-U6-gRNA carrier (structure such as figure 1 ) was digested with BsaI, and the enzyme digestion system was as follows:

[0047]

[0048]

[0049] 2. Gel recovery after enzyme digestion of the carrier

[0050] 3. Preparation of CXCL12-gRNA fragment PCR

[0051] The upstream and downstream primers are annealed, wherein the sequence of the forward primer is shown in SEQ ID NO.2, and the sequence of the reverse primer is shown in SEQ ID NO.3.

[0052] Table 2

[0053]

[0054] Use a PCR instrument or a water bath to mix the above components, incubate at 95°C for 4 minutes, then take it out and place it at room temperature to cool

[0055] 3. The processed target fragment and carrier ligation reaction system (20μl):

[0056] table 3

[0057]

[0058] The above reaction system is placed in a warm bath at 22°C for 20 minutes. If the follow-up operation cannot be performed immediately after the reaction is completed, the reaction sample can ...

Embodiment 2

[0063] Injection of adeno-associated virus recombinant vector for CXCL12 knockout into glioma model mice inhibits tumor growth and angiogenesis in mice

[0064] 1. Experimental materials

[0065] Nude mice aged 4-6 weeks (the mice can be purchased from Shanghai Slack Experimental Animal Co., Ltd.), weighing 16-20 g.

[0066] 2. Experimental method

[0067] 2.1 Mouse tumor modeling

[0068] Each of 24 nude mice was subcutaneously inoculated with 1*10^8 U87 cells in the anterior axilla. After 2 weeks, 24 mice were randomly divided into 3 groups, PBS group (blank control group), AAV-GFP group and AAV-CXCL12-KO group. The growth of nude mice was observed every day.

[0069] 2.2 Injection of AAV-GFP group and AAV-CXCL12-KO, control adeno-associated virus and PBS group

[0070] Aspirate 5ul of liquid and insert the needle at a distance of 5mm from the tumor with a microinjection needle, move subcutaneously to the glioma site, and inject slowly. Inject again every 2 days. The ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The present invention provides an adeno-associated virus recombinant vector for knocking out the CXCL12 gene. The recombinant vector comprises at least the following operably linked sequence elements:5 '-terminal inverse repeat sequence, CMV promoter, sacas9 sequence, polyA signal sequence, U6 promoter sequence, gRNA sequence, 3'-terminal inverse repeat sequence. CXCL12 gene knockout vector can effectively inhibit tumor growth and tumor angiogenesis of U87 cell glioma model, and can effectively inhibit tumor growth and tumor angiogenesis of U87 cell subcutaneous glioma model in nude mice. Thetechnical proposal of the invention can be applied to the treatment of glioma diseases.

Description

technical field [0001] The invention relates to an adeno-associated virus recombinant vector for knocking out CXCL12 gene, its construction method and application, and belongs to the field of biotechnology. Background technique [0002] CXCL12, also known as stromal cell-derived factor-1 (SDF-1), is a small molecule cytokine that belongs to the chemokine protein family. CXCL12 and its receptor (CXCR4) play an important role in the occurrence and development of many physiology and pathology. In tumors, intracellular signals are changed through different pathways, which can promote the proliferation, movement, adhesion and invasion of tumor cells. Affect the occurrence, development and prognosis of tumors. [0003] Adeno-associated virus (adeno-associated virus, AAV) is a genus of dependent viruses in the Parvoviridae family. The size of the virus particle is about 20-26 nm. The complete life cycle requires the participation of helper virus (usually adeno-associated virus or ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N15/864C12N5/10C12N15/66A61K35/768A61P35/00
CPCA61P35/00A61K35/768C12N15/66C12N15/86C12N2750/14143Y02A50/30
Inventor 陈意雄蔡晓龙邹杰朱鹏飞白易鑫
Owner 汉恒生物科技(上海)有限公司
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com