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

AmiRNA-HIF-1alpha sequence containing recombinant plasmid

A technology of HIF-1 and amirna-hif-1, applied in the field of recombinant plasmids, can solve problems such as disputes over the independent role of HIF family members

Inactive Publication Date: 2014-11-05
SHANGHAI PUTUO DISTRICT CENT HOSPITAL
View PDF2 Cites 4 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The HIF signaling system is responsible for the regulation of VEGF and tumor angiogenesis, however the independent role of HIF family members in this process is still controversial

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
  • AmiRNA-HIF-1alpha sequence containing recombinant plasmid
  • AmiRNA-HIF-1alpha sequence containing recombinant plasmid
  • AmiRNA-HIF-1alpha sequence containing recombinant plasmid

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Example 1: pcDNA TM 6.2 Construction of recombinant plasmid of GW / EmGFP-amiRNA-HIF-1α (abbreviated as p-amiRNA-HIF-1α).

[0038] 1. The design, synthesis and functional verification process of HIF-1α siRNA is as follows: figure 1 shown, including the following steps:

[0039] 1.1 Using HIF-1α CDS as a template, design its siRNA sequence, following the principles of complete complementary pairing with the template and not interfering with the expression of other genes, and design its control sequence, which is synthesized by takala company. And carry out 2-MO modification at the 5' end.

[0040] Target siRNA sequence: 5'-UGUGAGUUCGCAUCUUGAU-3'

[0041] Control siRNA sequence: 5'-UACACCGUUAGCAGACACC-3'

[0042] 1.2 Culture in McCoy’s 5A complete medium containing 10% newborn bovine serum, 100U / ml penicillin, 100μg / ml streptomycin (37°C 5% CO 2 , saturated humidity) human colorectal carcinoma HTC-116 cell line, press 1×10 6 Seed cells in exponential growth phase in 2...

Embodiment 2

[0090] Example 2: Human colorectal cancer cells were transfected with amiRNA-HIF-1α plasmid, and cell proliferation was observed.

[0091] Cultured in McCoy’s 5A complete medium containing 10% newborn calf serum, 100 U / ml penicillin, 100 μg / ml streptomycin (37°C, 5% CO 2 , saturated humidity) human colorectal carcinoma HTC-116 cell line, press 1×10 6 Seed cells in exponential growth phase in 24-well plates at 37°C in 5% CO 2 Incubate overnight in an incubator until the cell density reaches 60%-80%.

[0092] The above conventionally cultured human colorectal cancer HCT-116 cells were randomly divided into blank group, vector plasmid group, and recombinant plasmid group, and different plasmids were added according to the following doses for transient transfection:

[0093] (1) Blank group: normal HCT-116 cells, n=3;

[0094] (2) Vector plasmid set: p-amiR, n=3;

[0095] (3) Recombinant plasmid group: p-amiRNA-HIF-1α, n=3.

[0096] In a 1.5ml centrifuge tube, prepare the com...

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 an amiRNA-HIF-1alpha (artificial microRNA-hypoxia-inducible factor-1alpha) sequence containing recombinant plasmid, which contains a coding sequence of amiRNA-HIF-1alpha and a vector plasmid DNA sequence. The vector plasmid DNA sequence contains a regulatory sequence in connection with the amiRNA-HIF-1alpha, and the vector plasmid is a eukaryotic expression plasmid able to replicate and survive in a host. Construction of the recombinant plasmid includes synthesis of the amiRNA-HIF-1alpha sequence, connection of a target fragment and the vector plasmid, transformation of escherichia coli competent cells by the connection product, electrophoresis identification of the recombinant plasmid and sequencing, and other steps. The recombinant plasmid provided by the invention can be used for studying the action mechanism of amiRNA-HIF-1alpha in tumor angiogenesis, proliferation and apoptosis, invasion and metastasis, and development and production of amiRNA-HIF-1alpha based antitumor drugs, thus providing a new way for prevention and treatment of malignant tumors.

Description

technical field [0001] The invention relates to a recombinant plasmid, in particular to a recombinant plasmid containing amiRNA-HIF-1α. Background technique [0002] Tumor angiogenesis or angiogenesis (Vascμlarization) is a very critical factor for the growth of solid tumors. The growth, invasion and metastasis of malignant tumors depend on the formation of tumor blood vessels. Without angiogenesis, the growth of primary tumors will not exceed 1 ~2mm 3 , its continued growth must rely on the formation of new blood vessels to provide sufficient nutrients and oxygen, and take away metabolites. At the same time, the walls of new blood vessels are incomplete and in a state of high permeability, which is conducive to the metastasis of tumor cells. Tumor angiogenesis is a very complex process involving the interaction of tumor cells, endothelial cells, phagocytes, and their secreted factors, which may be promoters of tumor angiogenesis (eg, VEGF, Ang-2, bFGF, IL -8, etc.) or inh...

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): C12N15/85C12N15/66A61P35/00
Inventor 殷佩浩彭文邱艳艳奉典旭汤庆丰胡送娇梁波包益洁石晓静于卉闫翠娜冯雯陈亚峰郭晓玲邹瑜许婕
Owner SHANGHAI PUTUO DISTRICT CENT HOSPITAL
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