FoMV-mediated GFP-ATG8 expression vector and application thereof

A GFP-ATG8, expression vector technology, applied in the biological field, can solve problems such as small gene fragments

Pending Publication Date: 2021-03-02
TIANJIN NORMAL UNIVERSITY
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  • Abstract
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AI Technical Summary

Problems solved by technology

BSMV can also be applied to VOX, but the gene fragments that its vector can stably carry are small (less than 450-500 bp), and the target gene can only be combined with the virus gamma Expressed in the form of b gene fusion

Method used

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  • FoMV-mediated GFP-ATG8 expression vector and application thereof
  • FoMV-mediated GFP-ATG8 expression vector and application thereof
  • FoMV-mediated GFP-ATG8 expression vector and application thereof

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Embodiment 1

[0044] FoMV virus-mediated GFP-ATG8 expression vector for monitoring wheat autophagy after starvation and drug treatment

[0045] First construct the GFP-TaATG8a expression vector mediated by the FoMV virus with the method of the present invention for monitoring the autophagy level of wheat after starvation and drug treatment, including the following steps:

[0046] 1. Nicotiana benthamiana ( Nicotiana benthamiana ) and routine cultivation of wheat seedlings. benthamiana ( Nicotiana benthamiana ) and wheat variety Yangmai 158 were sown in pots filled with nutrient soil and vermiculite, and placed in a light incubator at 23°C, 16 h light / 8 h dark. The leaves of Nicotiana benthamiana plants aged 4-5 weeks are used for agricultural stems

[0047] fungus agroinfiltration, two-leaf stage wheat seedlings were used for virus inoculation.

[0048] 2. Construction of VOX vector of GFP-TaATG8a fusion gene. Use the VOX vector based on FoMV to construct the VOX vector of GFP-TaAT...

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Abstract

The invention constructs a FoMV-mediated GFP-ATG8 expression vector and application thereof. A method takes TaATG8a as a target for monitoring autophagy, a VOX technology based on FoMV is adopted forexpressing GFP-TaATG8a fusion protein on a wheat plant, the expression situation and subcellular localization situation of the fusion protein in leaves and root tissues are observed through fluorescence, and a technical platform for monitoring the autophagy level on a wheat living plant with overexpression GFPTaATG8 as the target is established. The platform is successfully applied to observationof representation autophagosomes and autophagovesicles in leaf epidermis, mesophyll cells and root cells under hunger and drug treatment, and the invention shows that the vector can be applied to autophagy level monitoring and autophagy function research in the wheat growth and development process or the environmental factor response process. Research results provide technical support for deep exploration of autophagy regulation and control mechanisms and physiological functions of important crop wheat.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to a FoMV virus-mediated GFP-TaATG8a expression vector and its application in monitoring the autophagy level of wheat. Background technique [0002] Autophagy (autophagy for short) is closely related to plant growth, development, aging, cell death and stress response. Monitoring the level of autophagy in cells is a necessary means to carry out research on the regulation mechanism and physiological function of autophagy. In the process of autophagy, the substrate to be degraded in the cytoplasm is wrapped into the autophagosome with a double-membrane structure. With the fusion of the outer membrane of the autophagosome and the membrane of the vacuole (lysosome in animals), the inner membrane and the wrapped autophagosome Substrates (now called autophagic vesicles) enter the vacuolar lumen to be broken down. A variety of autophagy-related (autophagy-related, ATG) factors are involved in t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/83C07K14/415A01H5/00A01H6/46G01N21/64
CPCC12N15/8203C12N15/8212C12N9/63G01N21/6486
Inventor 王华忠岳洁瑜胡蕊洁
Owner TIANJIN NORMAL UNIVERSITY
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