Adeno-associated virus expression vector and construction, virus preparation and cell marking method thereof

Abstract

The invention discloses a construction method of an adeno-associated virus expression vector, and the method comprises the following steps: obtaining a first gene sequence and a second gene sequence, the first gene sequence comprising a nuclear localization protein gene sequence, a chromogenic protein gene sequence and a receptor protein gene sequence; introducing an enzyme digestion site into the first gene sequence, and performing enzyme digestion treatment on the first gene sequence and the second gene sequence; wherein the second gene sequence carries a restriction enzyme cutting site; and connecting the first gene sequence and the second gene sequence after enzyme digestion treatment to obtain the adeno-associated virus expression vector. The invention also discloses a preparation method of the adeno-associated virus, a cell marking method and an expression vector. In this way, marking and positioning of whole-brain specific type neurons in a specific brain region can be achieved.

Description

technical field [0001] This application relates to the field of genetic engineering, in particular to an adeno-associated virus expression vector and its construction, virus preparation, and cell labeling methods. Background technique [0002] The brain is the most important central mechanism of the human body, and the neural network is the material basis of the brain's work. Abnormalities of the neural network often lead to various neurological diseases. Efficient retrograde labeling tools Viral vectors can manipulate upstream neural networks transmitted to specific brain regions for imaging, which is critical in the elucidation of neural circuit structure and function. The known efficient retrograde labeling tool viruses mainly include AAV2-Retro (Neuron, 2016, 92(2): 372-382.) and RV-ΔG (N2cG) (NeurosciBull, 2020, 36(3): 202-216.) . However, AAV2-Retro has brain region selectivity, and it mainly infects cortical neurons. Although RV-ΔG (N2cG) can retrogradely label neur...

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Problems solved by technology

However, AAV2-Retro has brain region selectivity, and it mainly infects cortical neurons. Although RV-ΔG (N2cG) can retrogradely label neural networks in a broad spectrum, it cannot achieve the specificity of retrogradely labeled neurons.

Known efficient retrograde labeling tool viruses cannot achieve whole-brain retrograde labeling of specific types of neurons for specific brain region reception

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