Agrobacterium-mediated orychophragmus violaceus transgene method

An Agrobacterium-mediated, transgenic technology, applied in genetic engineering, plant genetic improvement, botanical equipment and methods, etc., can solve the problems of long experimental period and low transformation efficiency, and achieve short experimental period, high transformation efficiency and large scale. The effect of practical application value

Inactive Publication Date: 2015-08-05
INST OF BOTANY JIANGSU PROVINCE & CHINESE ACADEMY OF SCI
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  • Abstract
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  • Application Information

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Problems solved by technology

[0008] The invention provides an Agrobacterium-mediated transgenic method of Zhugecai. Through the spraying method of flowers, the inflorescence of Zhugecai is contacted with Agrobacterium under living conditions to complete the transformation of genetic cells, and obtain the transformation by screening the offspring. Plants, which solve the problems of low transformation efficiency and long experiment period of Zhuge vegetable by conventional tissue culture transformation method

Method used

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  • Agrobacterium-mediated orychophragmus violaceus transgene method

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Embodiment 1

[0028] (1) Zhuge vegetable planting

[0029] Sow the seeds of Zhugecai on February 20th in the seedbed for seedling cultivation. The soil of the seedbed should be fertile and moist, covered with wheat straw or plastic film. About one month after the seedlings have emerged, ventilate properly. When the height of the seedlings is 3-4cm, gradually remove the cover to allow the seedlings to receive direct light. When the height of the seedlings is 8-10cm, choose cloudy days or before the rain for transplanting. 10-15cm, pour water to fix the roots immediately after planting. Plant 1000 Zhuge vegetables, covering an area of ​​about 50㎡ ( figure 1 ). The fertilizer and water management of Zhuge vegetable is carried out according to the conventional method, and it blooms from April to May of that year. At this point, choose the best time to transform Agrobacterium zhugeli.

[0030] (2) Preparation of Agrobacterium containing herbicide-resistant Basta selection marker gene plasmi...

Embodiment 2

[0045](1) Zhuge vegetable planting

[0046] Sow the seeds of Zhugecai on February 20th in the seedbed for seedling cultivation. The soil of the seedbed should be fertile and moist, covered with wheat straw or plastic film. About one month after the seedlings have emerged, ventilate properly. When the height of the seedlings is 3-4cm, gradually remove the cover to allow the seedlings to receive direct light. When the height of the seedlings is 8-10cm, choose cloudy days or before the rain for transplanting. 10-15cm, pour water to fix the roots immediately after planting. Plant 1,000 Zhuge vegetables, covering an area of ​​about 50 square meters. The fertilizer and water management of Zhuge vegetable is carried out according to the conventional method, and it blooms from April to May of that year. At this point, choose the best time to transform Agrobacterium zhugeli.

[0047] (2) Preparation of Agrobacterium containing herbicide-resistant Basta selection marker gene plasmid ...

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Abstract

The present invention discloses an agrobacterium-mediated orychophragmus violaceus transgene method. According to the method, when buds appear on the main branch and the partial side branch of the orychophragmus violaceus and several flowers flower, the flowering flowers on the main inflorescence and each branch inflorescence are removed, and only the buds are retained; agrobacterium containing an expression vector is suspended with an osmotic culture medium and then is sprayed on the orychophragmus violaceus inflorescence, and wrapping is immediately performed with parchment paper after infiltration, wherein the osmotic culture medium contains a surfactant Silwet L-77 and sucrose, the optimal concentration of the Silwet L-77 is 500-1000 [mu]l/L, and the optimal concentration of the sucrose is 30-60 g/L; the same inflorescence is subjected to continuous spraying three times, and the interval time is 2-3 days; and 7-10 days after the conversion, the parchment paper bag on the inflorescence, and the flowers and the buds on the top end of the inflorescence are picked out. According to the present invention, the operation of the method is simple, the required cost is low, the transgene efficiency is high, the experimental period is short, and the good application prospects are provided in the field of the utilization of genetic engineering to cultivate novel special oil crops and the establishment of the orychophragmus violaceus mutant library.

Description

technical field [0001] The invention belongs to the field of plant genetic engineering, and relates to an Agrobacterium-mediated transgenic method for Zhuge vegetable. Background technique [0002] Zhuge Cai ( Orychophragmus violaceus (L.) O. E. Schulz) is a plant of the Brassica family of Brassicaceae. The oil content of the seeds is as high as 35.8%. It is rich in linoleic acid, oleic acid, palmitic acid, etc. that are beneficial to the human body, while linolenic acid is harmful to the human body. , erucic acid content is very low, it is a new type of high-quality oil plant, and it is also an excellent provenance for the creation and breeding of new varieties of special oil crops. Zhuge Cai grows wild in plains, mountains and roadsides. Wide distribution, strong adaptability, easy cultivation and management, and good prospects for development and utilization. [0003] At present, transgenic technology is developing rapidly and has comprehensively promoted the upgradin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/84A01H5/00
Inventor 孙小芹杭悦宇张艳梅
Owner INST OF BOTANY JIANGSU PROVINCE & CHINESE ACADEMY OF SCI
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