Method for assisting recombinant rabies virus in packaging rAAV virus with efficient reverse cross-single-grade synaptic nerve cells and application of rAAV virus

Abstract

The invention discloses a method for assisting a recombinant rabies virus in packaging a recombinant adeno-associated virus with efficient reverse cross-single-grade synapsis synaptic nerve cells andapplication. The method comprises the steps that an expression plasmid carried with a fluorescent protein gene and a TVA receptor gene and an expression plasmid carried with a rabies virus membrane protein (G protein) gene are pre-mixed into a plasmid mixture, then the plasmid mixture and a recombinant adeno-associated virus assistant packaging plasmid are used for cotransfection and cell packaging, and through purification, a single recombinant adeno-associated virus set with good stability is obtained. Compared with a method that different expression plasmids are packaged into the recombinant adeno-associated virus separately and then mixed infection is caused, the method has the advantages that the rejection problem caused by infection after mixing of different recombinant adeno-associated viruses can be obviously solved, the obtained novel recombinant adeno-associated virus can assist the recombinant rabies virus in more efficiently marking a single-grade input network of brain nerve cells in the full brain, and that is to say, the full-brain single-grain input network with more complete nerve cells of a specific brain area of the brain can be obtained.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a packaging method and application thereof for assisting recombinant rabies virus to cross single-level synapses efficiently and reversely in nerve cells. Background technique [0002] A neural circuit is composed of a very large number and a variety of nerve cells connected in a certain pattern, and is the structural basis for the brain to perform various functions. The precise resolution of the structural connections of neural circuits will help people better understand how the brain performs different functions. Recombinant rabies virus reverse single-level synaptic labeling system has been widely used in the study of neural circuits as a tool for analyzing the reverse single-level input network of nerve cells. [0003] Recombinant rabies virus reverse cross-synaptic labeling system At present, recombinant adeno-associated virus (rAAV) is commonly used as a helper vir...

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Problems solved by technology

Therefore, the three proteins (TVA, XFP, RG) in the above two rAAV helper virus mixing methods cannot be completely and efficiently expressed, which leads to the inability of the recombinant RV virus to efficiently label the whole brain input network of neurons in the target brain area. Therefore, there are two kinds of rAAV There is still a certain gap between the whole-brain single-level input network and all the single-level input networks of nerve cells that are mixed with helper virus injection and combined with recombinant RV virus labeling, which restricts people's understanding of the complete neural circuit organization structure of nerve cells in specific brain regions

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