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Efficient betula platyphylla root transgenic method

A transgenic, birch technology, applied in horticultural methods, genetic engineering, plant genetic improvement and other directions, can solve the problems of low efficiency, long transformation period, long explant culture period, etc., achieves simple operation, shortened transformation period, shortened culture effect of cycles

Pending Publication Date: 2022-08-02
NORTHEAST FORESTRY UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to solve the problems of long explant culture period, long transformation period and low efficiency in the existing white birch transgenic system, the present invention provides a highly efficient method for transgenic white birch root

Method used

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  • Efficient betula platyphylla root transgenic method
  • Efficient betula platyphylla root transgenic method
  • Efficient betula platyphylla root transgenic method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] The present embodiment provides an efficient method for transgenic birch root, comprising the following steps:

[0034] Step 1. Micro-cuttings of wild-type tissue-cultured birch seedlings are placed in rooting medium for rooting culture at room temperature.

[0035] The rooting medium was prepared by adding 20g / L sucrose, 0.2mg / L NAA, 0.4g / L carbon powder and 6g / L agar on the basis of 1 / 2MS medium, and the pH was 5.8-6.2.

[0036] Step 2. Place the Agrobacterium containing the pBI121-GUS vector of the 35S promoter driving the expression of the GUS gene in 5 mL of liquid LB medium containing 20 mg / L rifampicin and 50 mg / L kanamycin. Incubate at 220rpm for 2-3d.

[0037] The Agrobacterium competent cells EHA105 and the pBI121-GUS vector used in this example were purchased commercially.

[0038] Step 3. Transfer 500 μL of the Agrobacterium culture solution from step 2 to a new 20 mL double-antibody LB medium and continue to culture to OD 600 For 0.6-0.8, the cells were ...

Embodiment 2

[0053] The present embodiment provides an efficient method for transgenic birch root, comprising the following steps:

[0054] Step 1. Micro-cuttings of wild-type tissue-cultured birch seedlings are placed in rooting medium for rooting culture at room temperature.

[0055] The rooting medium was prepared by adding 20g / L sucrose, 0.2mg / L NAA, 0.4g / L carbon powder and 6g / L agar on the basis of 1 / 2MS medium, and the pH was 5.8-6.2.

[0056] Step 2. The Agrobacterium containing the pROKII-LUC vector was placed in 5 mL of liquid LB medium containing 20 mg / L rifampicin and 50 mg / L kanamycin, and cultured at 28° C. and 220 rpm for 2-3 days.

[0057] The Agrobacterium competent cell EHA105 used in this example was purchased commercially, and the pROKII-LUC vector was constructed by Wang Chao's team from the Forest Genetics and Breeding Laboratory of Northeast Forestry University according to conventional gene ligation methods in the field.

[0058] Step 3. Transfer 500 μL of the Agro...

Embodiment 3

[0071] The present embodiment provides an efficient method for transgenic birch root, comprising the following steps:

[0072] Step 1. Micro-cuttings of wild-type tissue-cultured birch seedlings are placed in rooting medium for rooting culture at room temperature.

[0073] The rooting medium was prepared by adding 20g / L sucrose, 0.2mg / L NAA, 0.4g / L carbon powder and 6g / L agar on the basis of 1 / 2MS medium, and the pH was 5.8-6.2.

[0074] Step 2. Place the Agrobacterium containing the pEgP237-FLA9-2A-GFP vector in 5 mL of liquid LB medium containing 20 mg / L rifampicin and 50 mg / L kanamycin, and culture at 28°C at 220 rpm for 2- 3d.

[0075] The Agrobacterium competent cell EHA105 used in this example was purchased commercially, and the pEgP237-FLA9-2A-GFP vector was constructed by Wang Chao's team from the Forest Genetics and Breeding Laboratory of Northeast Forestry University according to conventional gene ligation methods in the field.

[0076] Step 3. Transfer 500 μL of t...

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Abstract

The invention relates to an efficient betula platyphylla root transgenosis method, and belongs to the technical field of plant transgenosis. In order to solve the problems of long culture period, long transformation period and low efficiency of explants of an existing betula platyphylla transgenosis system, the invention provides an efficient betula platyphylla root transgenosis method which comprises the following steps: selecting betula platyphylla roots with root tiller reproductive capacity as a transgenosis material; the method comprises the following steps: culture of white birch roots, genetic transformation of agrobacterium carrying a target gene, induction of callus formation through co-culture, induction of adventitious bud formation through resistance screening, and finally formation of a complete plant through induction. Compared with an existing betula platyphylla transgenosis method, the method has the advantages that the transformation period is shortened, the transformation efficiency is improved, the difficulty in explant selection is avoided, the genetic background of transgenic offspring plants is unified, the utilization rate of materials is improved, and the problem of difficulty in degerming is further solved. The efficient betula platyphylla root transgenic method provided by the invention is simple to operate and easy to master.

Description

technical field [0001] The invention belongs to the technical field of plant transgenesis, in particular to an efficient method for transgenic birch roots. Background technique [0002] White birch (Betula platyphylla Suk.) is a birch family birch and is a deciduous tree. It grows fast and is a pioneer tree species of secondary forests in Northeast China. It has strong cold resistance and prefers acidic soil. It is used in plywood, veneer furniture manufacturing and pulp materials has a wide range of applications. Because of its important ecological, ornamental and practical economic value, it has always been listed as one of the important tree species studied by the national science and technology plan. The scope of its genetic improvement is also expanding, and its main goal is to cultivate new varieties of fast-growing, high-quality and high-resistance trees. Therefore, the use of molecular biology methods to provide materials for the cultivation of new varieties of for...

Claims

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Application Information

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IPC IPC(8): C12N15/84A01H5/06A01H6/00A01H4/00
CPCC12N15/8205A01H4/002Y02P60/40
Inventor 于颖王超石晶静高岩张嘉薇王荣国苏丽红
Owner NORTHEAST FORESTRY UNIVERSITY
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