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A method for observing long-distance transport of fusion proteins to rice leaf buds

A fusion protein and long-distance technology, applied in the preparation of test samples, measuring devices, instruments, etc., to achieve the effect of simple methods

Inactive Publication Date: 2015-09-02
YUNNAN AGRICULTURAL UNIVERSITY
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Problems solved by technology

The transport of Magnaporthe oryzae effector proteins in rice root tissues has been reported, but there is no report on the transport of Magnaporthe oryzae proteins in rice plants, that is, from rice root tips to leaf buds

Method used

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Embodiment Construction

[0016] In order to make the object, technical solution and advantages of the present invention more clear, the present invention will be further described in detail below in conjunction with the examples. It should be understood that the specific embodiments described here are only used to explain the present invention, not to limit the present invention.

[0017] 5.0 mg / ml of GFP fusion protein treated seeds germinated to 14 days old rice young plants at the root tip of 0.5cm for 24 hours, rinsed in sterilized water for 4 hours, leaf buds were frozen and sectioned, PI stained the sections, and GFP was directly observed with a confocal microscope Transport of fusion proteins in rice plants.

[0018] The expression product of the effector protein gene of Magnaporthe grisea was collected by centrifugation at 4°C, and the cells were suspended with freshly prepared buffer solution (20mM Tris-HCl, pH7.4; 200mM NaCl; 1mM EDTA; 10mM β-mercaptoethanol). The supernatant was collected ...

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Abstract

Disclosed is a method for observing long-distance transport of fusion protein to the leaf bud of oryza sativa, the method comprising: using GFP fusion protein to treat the root tip of the oryza sativa, freezing and sectioning the leaf bud of an oryza sativa plant having the root tip treated, dying a leaf bud section by using PI fluorescent dye, and using a confocal microscope to directly observe the transport of the GFP fusion protein from the root tip to the leaf bud of the oryza sativa. The method uses the GFP fusion protein to treat the root tip of the oryza sativa for a length of 0.5 cm, freezes and sections the leaf bud, dyes a leaf bud section, and uses a confocal microscope to observe the transport of the GFP fusion protein from the root tip to the leaf bud of the oryza sativa, thus allowing learning directly and qualitatively about the long-distance transport condition of the GFP fusion protein within the oryza sativa plant, and providing convincing evidence for the ability of effector protein to induce the system defense response of the oryza sativa. In addition, the method is easy, accurate, quick, and of important usage value.

Description

technical field [0001] The present invention relates to a method of using GFP fusion protein to treat rice root tips to observe the long-distance transport of the fusion protein to rice leaf buds, and in particular to a method for treating rice root tips at 0.5 cm by using the prokaryotic expression product of the rice blast fungus protein gene fused with GFP, The invention discloses a laser confocal microscopy method for observing the translocation of GFP fusion protein from root tip 0.5cm to rice leaf buds, belonging to the fields of plant protection and cell biology. Background technique [0002] Pathogenic bacteria transport effectors through the type III secretion system, and its transport mechanism has been studied relatively clearly. But for filamentous fungi, how effector proteins are transported into host cells remains unknown, although specialized infection structures such as haustoria are involved in the transport of effector proteins. Oomycete effector proteins ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64G01N1/30
CPCG01N21/6458
Inventor 杨静李成云刘林朱有勇杜云龙徐畅施竹凤
Owner YUNNAN AGRICULTURAL UNIVERSITY
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