Preparation method and applications of humanized gene modified animal models

A humanized, non-human animal technology, applied in the fields of botanical equipment and methods, biochemical equipment and methods, plant genetic improvement, etc., can solve the problems of poor cell survival, short lifespan, and reduced CD4+ T cell proliferation, etc. To save time and cost, speed up the research and development process, and reduce the risk of drug development

Active Publication Date: 2018-08-21
BIOCYTOGEN PHARMACEUTICALS (BEIJING) CO LTD +1
View PDF2 Cites 17 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It was found that DC cells in mice lacking CD137 can mature normally, but after maturation, the cell survival is poor and the life span is short; and it is found that in vivo (in vivo) due to the lack

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
  • Preparation method and applications of humanized gene modified animal models
  • Preparation method and applications of humanized gene modified animal models
  • Preparation method and applications of humanized gene modified animal models

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0170] Example 1 Homologous Recombination Fragment Primer Design and PCR Amplification

[0171] According to the experimental design scheme, primers for amplifying seven homologous recombination fragments (A1, A2-1, A2-2, A3, B, C1, C2) were designed. The primer sequences are shown in Table 1.

[0172] Table 1 Homologous recombination fragments and their corresponding lengths and primer sequences

[0173]

[0174]

[0175] Using BAC as a template, 7 homologous recombination fragments were amplified using KOD-plus-polymerase. Among them, A1, A3, B, C1, and C2 were selected from Bacterial Artificial Chromosome (BAC) containing the mouse CD137 genome (product number: RPCI23.C, clone ID: 448J14, hereinafter referred to as "bacteria containing mouse BAC") As the template, A2-1 and A2-2 selected the bacterial artificial chromosome containing the human CD137 genome (article number: RPCI11.C, clone ID: 208A7, hereinafter referred to as "bacteria containing human BAC") as the te...

Embodiment 2

[0179] Example 2 Construction of Homologous Recombination Targeting Vector

[0180] Schematic diagram of comparison between mouse Cd137 gene and human CD137 gene figure 1 shown. The inventor designed figure 2 The targeting strategy shown, figure 2 The design of the targeting vector is also shown. Mouse Cd137 (based on NM_011612.2, ie SEQ ID NO: 15 and its corresponding protein expression NP_035742.1, ie SEQ ID NO: 16) has a total of 8 exons, of which exon 1 does not code, and exon 2 Some are not coded. The present inventors used human CD137 gene (based on NM_001561.5, namely SEQ ID NO: 17 and its corresponding protein expression NP_001552.2, namely SEQ ID NO: 18) for the main part of exon No. 2-7 of mouse Cd137 gene Fragment replacement, and adding neo gene to the targeting vector for positive clone screening. The length of its 5' homology arm (SEQ ID NO:19) is 4847bp, the length of its 3' homology arm is 4728bp (SEQ ID NO:20), and the length of the human DNA fragment ...

Embodiment 3

[0199] Example 3 Verification of carrier pDTA-down-ABC

[0200] Randomly select 4 pDTA-down-ABC clones, and use 3 groups of restriction endonucleases for digestion verification. Among them, 9010bp+5834bp+4165bp+3058bp+1448bp+494bp should appear when using XhoI+NdeI digestion, and using SalI+SmaI Enzyme digestion should produce 11641bp+7563bp+4083bp+722bp, and digestion with BstZ17I+KpnI should produce 12111bp+7897bp+3582bp+419bp. The results of enzyme digestion showed that plasmids 1, 2, and 3 were all in line with expectations, see Figure 5 , Plasmids 1 and 2 were sent to the sequencing company for sequencing and confirmed to be correct.

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 relates to humanized gene modified non-human animals, especially genetically modified rodents, and especially genetically modified mice, and more specifically relates to a construction method of humanized CD137 gene animal models, and applications of the humanized CD137 gene animal models in biomedicine field.

Description

technical field [0001] This application relates to the establishment method and application of a humanized genetically modified animal model, in particular, to the construction method of a humanized CD137 genetically modified animal model and its application in biomedicine. Background technique [0002] Experimental animal disease models are indispensable research tools for the study of the etiology and pathogenesis of human diseases, the development of prevention technologies and the development of drugs. However, due to the differences in the physiological structure and metabolic system between animals and humans, traditional animal models cannot well reflect the real conditions of the human body. It is an urgent need for the biomedical industry to establish disease models in animals that are closer to the physiological characteristics of humans . [0003] With the continuous development and maturity of genetic engineering technology, the replacement or replacement of hom...

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): C12N15/85C12N5/10C12N15/12C12N15/62C07K19/00A01K67/027
CPCA01K67/0278A01K2217/072A01K2227/105A01K2267/03A01K2267/0331A01K2267/0387C07K14/70578C07K2319/00C12N15/85A61K49/0008C12N15/8509C12N2015/8572
Inventor 沈月雷郭雅南白阳赵磊黄蕤姚佳维张美玲
Owner BIOCYTOGEN PHARMACEUTICALS (BEIJING) CO LTD
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

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

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap