Expression vector for stably indicating cell autophagy activities, establishing method and application method thereof

A technology for expressing vectors and cells, which is applied in the field of biomedicine, can solve problems such as the distortion of fluorescent spot observation, difficulty in screening stable strains and amplification, increase the difference in biological characteristics between transfected cells and mother cells, and achieve quantitative analysis of autophagy The method is simple and easy to use, the indication of autophagy is sensitive and reliable, and the effects on a wide range of cell types
CN101892265AInactive Publication Date: 2010-11-24ZHONGSHAN HOSPITAL FUDAN UNIV
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Patent Information

Authority / Receiving Office
CN · China
Current Assignee / Owner
ZHONGSHAN HOSPITAL FUDAN UNIV
Publication Date
2010-11-24
Estimated Expiration
Not applicable · inactive patent

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Abstract

The invention belongs to the biomedicine field, and aims at providing an expression vector for stably indicating cell autophagy activities, an establishing method and an application method thereof. By constructing a slow-virus expression plasmid containing an EGFP-LC3 autophagy reporter gene and adopting a four-plasmid system, the invention establishes a slow-virus expression vector capable of enabling cells to stably express an EGFP-LC3 gene. By using the expression vector to infect a host cell, the EGFP-LC3 gene can be stably integrated in a genome of the host cell, and the host cell can stably and efficiently express the EGFP-LC3 and indicate cell autophagy changes. A stable autophagy indicating cell can be established by using the expression vector, and the cell autophagy changes are quickly and quantitatively analyzed by an image analysis method based on Top-hat operators and morphological filtration. The invention has the characteristics of stable and reliable autophagy indication, real-time sensitivity, quick and accurate autophagy quantification, simply and convenient operation and the like, greatly facilitates autophagy observation and analysis, and can be widely used for autophagy-correlated researches.
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Description

technical field

[0001] The invention belongs to the technical field of biomedicine, and relates to an expression vector stably indicating cell autophagy activity and its establishment and application method. Background technique

[0002] Autophagy is a process of cell self-degradation and digestion, and is an important life activity of cells. Under normal physiological conditions, cells rely on autophagy to remove aging or damaged organelles and renew some important proteins; when cells are in a state of stress, autophagy activation is enhanced to maintain energy and metabolic homeostasis by recycling intracellular components , remove damaged proteins and organelles, limit oxidative damage, etc., are important mechanisms for maintaining cell homeostasis and survival. Because it is closely related to many human diseases, especially in the occurrence and development of major diseases such as tumors, it has become a research hotspot in recent years, and related research is inc...

Claims

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