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

Transgenic carrier system for promoting cell transplantation and gene expression and application thereof

A transgenic vector, gene expression technology, applied in the direction of plant gene improvement, application, vector, etc., can solve problems such as hindering application, uncommon target gene characteristics, tumor occurrence, etc., to avoid the influence of drug toxicity and achieve the effect of great application value

Inactive Publication Date: 2019-10-25
HANGZHOU DIANZI UNIV
View PDF3 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There are currently three types of in vivo screening methods: 1) The target gene has the ability to make the host cell continue to divide and promote survival, so that in vivo screening occurs naturally without any screening pressure, but the characteristics of this target gene are not common, That is, not all target genes have the ability to make the host cell continue to divide and promote survival; 2) In addition to the target gene, a screening gene is introduced at the same time, which enables the host cell to have drug-dependent ability to promote survival (so-called The cell growth switch cell growth switches), but this method shows that the screening only occurs at the level of short-term survival cells in large animals, so that it cannot maintain enough cells expressing the target gene for a long time, thus limiting its use; 3) except In addition to the target gene, a screening gene that resists screening pressure (usually toxic drugs) is introduced at the same time, so that the drug only eliminates non-genetically modified cells but not genetically modified cells, but this system hinders its application due to toxic drugs
4) When a gene with a growth advantage is used as a screening gene, it has the possibility of causing tumorigenesis
So far, no in vivo screening system that can overcome the above defects has been seen

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
  • Transgenic carrier system for promoting cell transplantation and gene expression and application thereof
  • Transgenic carrier system for promoting cell transplantation and gene expression and application thereof
  • Transgenic carrier system for promoting cell transplantation and gene expression and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1. In vivo screening system using BCL2 mutant S70A as screening gene

[0043] In Example 1, the BCL2 mutant S70A was used as the screening gene, and the retroviral vector MIGR1 was used as the vector. As mentioned above, S70A retains part of the anti-apoptotic ability of BCL2, but removes its tumor-promoting ability, so it is particularly suitable as a natural screening gene in vivo.

[0044] (1) Insert S70A before the IRES of MIGR1, which is the MIGR1-S70A plasmid, such as figure 1 shown.

[0045] (2) The retroviral supernatant was prepared with MIGR1-S70A plasmid and packaging cell BOSC23. Specifically, BOSC23 cells were cultured in 6-well cell culture plates in complete medium (Dulbecco's Modified Eagle's Medium, DMEM) complete medium (adding 10% fetal bovine serum, 100 units / ml penicillin, 100 μg / ml streptomycin, 2mM L-glutamine, etc.) (37°C, 5% CO 2 ), when the cells are 90% confluent, replace the old medium with fresh DMEM medium containing 5% FBS and ...

Embodiment 2

[0052] Example 2. In vivo screening system based on induction system and lentiviral vector using Bcl-2 mutant S70A as screening gene

[0053] Lentiviruses have safer properties than retroviruses. Therefore repeat the research of embodiment 1 with self-inactivating feline immunodeficiency virus ( image 3 , the empty viral vector was from SBI Company). The packaging cells used for virus production are 293T cells. In addition to the lentiviral vector, two other packaging plasmids (pFIV-34N, pVSV-G) should be transferred into the 293T cells. The lentivirus packaging method, bone marrow cell acquisition, virus infection and cell transplantation are the same as in Example 1.

[0054] Six months after transplantation, it was found that lentivirus-based S70A could increase the proportion of GFP-positive cells, but unlike retrovirus S70A, lentivirus S70A could only increase the proportion of GFP-positive cells to a low level, about 30%. By comparing the fluorescence intensity value...

Embodiment 3

[0055] Example 3. Non-myeloablative pre-transplant conditioning by 5-FU treatment

[0056] In clinical practice, transplant recipient patients often cannot use lethal doses of radiation to irradiate myeloablation, so it is very critical whether non-myeloablative pre-transplantation treatment with drugs is feasible. Before transplantation, the recipient mice were injected with 150 mg / kg body weight of pentafluorouracil (5-FU). Six months after transplantation, it was found that this pretreatment was sufficient to successfully transplant bone marrow cells infected by retrovirus MIGR1-S70A ( Figure 4 ), so this method is very suitable for clinical application.

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 invention discloses a transgenic carrier system for promoting cell transplantation and gene expression and application thereof. The system comprises a gene selecting system, a target gene expression system and a carrier used for carrying the two systems, wherein the gene selecting system is a system for silencing genes for inhibiting division or promoting cell death. Compared with the prior art, the system is not limited by whether the targeted gene has growth advantages or not; selecting is generated in the stem cell level and the differentiated cell level, and therefore, the quantity ofgenetically engineered cells can be stably kept or increased in the whole lifetime; selecting is conducted without drugs, and the influence of the drug toxicity is avoided; moreover, a mutant which only inhibits division or promotes cell death is employed, so that there is no risks of causing cancer.

Description

[0001] Original application date: 2016-10-08, original application number: 201610877184.9 [0002] Original patent title: A transgenic carrier system and its application for promoting cell transplantation and gene expression technical field [0003] The invention relates to a transgene carrier system, especially a carrier system capable of promoting cell transplantation and gene expression. Background technique [0004] Gene therapy or the establishment of adult (stem) cell-based animal bioreactors or animal models require stable target gene expression levels and stable target gene expression positive cell numbers in recipient patients or animals. However, in reality, it generally occurs that the efficiency of expressing the target gene in a single cell decreases and the proportion of cells expressing the target gene decreases. An approach to address the above deficiencies is to perform in vivo screening. There are currently three types of in vivo screening methods: 1) The...

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
Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/85C12N15/867
CPCC12N15/85C12N15/86C12N2800/30C12N2830/36C12N2740/15043C12N2740/11043
Inventor 王彦刈马珊
Owner HANGZHOU DIANZI UNIV
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