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Method for quickly separating vascular bundle, epidermis and mesophyll cells from plant leaves

A technology of plant leaves and mesophyll cells, which is applied in the field of plant separation, can solve problems such as difficult and difficult to avoid contamination of different tissues, and achieve high purity and speed up the effect of tissue-specific analysis

Pending Publication Date: 2019-09-20
国重科技(武汉)有限公司
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  • Abstract
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Problems solved by technology

But for transient expression, it is mainly used in superficial tissues, while deep tissues such as vascular bundle cells are more difficult
For stable transformation, it is difficult to avoid contamination of different tissues

Method used

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Experimental program
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Embodiment Construction

[0023] The present invention will be further described below in combination with specific embodiments.

[0024] The present invention proposes a method for rapidly separating vascular bundles, epidermis and mesophyll cells from plant leaves, comprising the steps of:

[0025] S1: Select plant seeds, use standard bleach to sterilize the surface of the seeds in a microcentrifuge tube, and sow the seeds with 0.5% (mass volume ratio) sucrose on the 0.5MS medium, and the distance between the seeds is about 0.7 mm to 1 cm. Store the seeds at 4°C for 1 day, and let the seeds wait for germination under normal growth conditions. The preparation method of MS medium is as follows:

[0026] Select 16 grams of agar powder and 10 grams of sucrose into 1,900 ml of distilled water, put them in a 2-liter bottle, adjust the pH value to 5.7-5.8 with 1 mole of potassium hydroxide per liter, add water until the volume of 2 liters, and obtain culture Base, pressurize the medium at 121°C for 20 minu...

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Abstract

The invention discloses a method for quickly separating vascular bundle, epidermis and mesophyll cells from plant leaves. The method includes following steps: S1, using a standard bleacher to disinfect the surfaces of seeds in a micro centrifugal tube; S2, transferring a culture medium in the S1 into a plant growth chamber for storage; S3, preparing a centrifugal tube of 1.5ml, and separating mesophyll protoplasm, wherein an enzyme solution of 1ml is placed in the centrifugal tube; S4, heating frozen enzyme liquid prepared in advance to 22-25 DEG C, and maintaining the same at room temperature in subsequent steps. The method is a separating technology scheme, and practices show that the method can separate out all three major leaf tissues (mesophyll, vascular bundle and epidermis) within 30min while purity is quite high.

Description

technical field [0001] The invention relates to the technical field of plant separation, in particular to a method for rapidly separating vascular bundles, epidermis and mesophyll cells from plant leaves. Background technique [0002] In the field of plant research, more and more experiments have shown that different tissues respond differently to the same environmental change. Therefore, it is the development direction of botany research to effectively isolate different plant tissues to test tissue-specific responses. Several methods have been developed to isolate specific cell types, including laser capture microdissection, the use of plants with tissue-specific green fluorescent protein expression to generate protoplasts, and activated fluorescent cell sorting. However, the method of laser capture microdissection is very complicated, and there are certain difficulties in processing a large number of samples. Another method is to measure promoter activity using luciferas...

Claims

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Application Information

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IPC IPC(8): C12N5/04
CPCC12N5/04C12N2509/00
Inventor 卢娜孔繁帆彭国杰
Owner 国重科技(武汉)有限公司
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