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Method for optimizing transgenosis efficiency of arabidopsis thaliana

A Arabidopsis thaliana, transgenic technology, applied in the field of Arabidopsis transgenic technology, can solve the problems of low transformation efficiency, low plant utilization rate, low transformation medium utilization rate, etc., to improve transformation efficiency, improve plant utilization rate, and prolong transformation the effect of time

Active Publication Date: 2018-09-14
GUANGXI BOTANICAL GARDEN OF MEDICINAL PLANTS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, this method has the following disadvantages: because only the robust plants of the first batch of bolting around the 4th week are selected as the dyeing material, and other plants with slightly longer growth periods are often discarded, and the utilization rate of the plants is low; Carry out dip-dyeing, prepare the transforming medium, that is, the dipping solution is generally 1-2 liters, and all the racemes that have removed pods and pollinated are soaked in the medium, and the transforming medium is no longer used after treatment, and the utilization rate of the transforming medium is low; This method obtains seeds, and only about 10 positive plants are obtained per 0.1 ml, and the transformation efficiency is low

Method used

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  • Method for optimizing transgenosis efficiency of arabidopsis thaliana
  • Method for optimizing transgenosis efficiency of arabidopsis thaliana

Examples

Experimental program
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Effect test

Embodiment 1

[0025] To optimize the transgenic efficiency of Arabidopsis thaliana, the inflorescences on the Arabidopsis thaliana plants cultivated for 4 weeks were soaked with the transformation medium, and each Arabidopsis thaliana plant was wrapped with plastic wrap, the plants were cultivated, and the seeds were collected.

[0026] Among them, the specific configuration method of the transformation medium is:

[0027] S1. Prepare YEP culture medium containing 50 mg / L rifampicin and 100 mg / L kanamycin, and mix 1 volume part of Agrobacterium containing target gene pBI121 vector with 500 volume parts of YEP culture medium , Activated and cultured in a constant temperature shaker at 160 rpm and 28°C for 18 hours, and mixed 1 volume part of the activated pBI121 vector containing the target gene with 100 vol parts of YEP culture medium at 160 rpm and 28°C. Cultivate for 12 hours on a constant temperature shaker at ℃ until the OD600 value is 1.8, and centrifuge to collect Agrobacterium;

[0028] S2...

Embodiment 2

[0033] To optimize the transgenic efficiency of Arabidopsis thaliana, the inflorescences on the Arabidopsis thaliana plants cultivated for 4 weeks were soaked with the transformation medium, and each Arabidopsis thaliana plant was wrapped with plastic wrap, the plants were cultivated, and the seeds were collected.

[0034] Among them, the specific configuration method of the transformation medium:

[0035] S1. Prepare YEP culture medium containing 50 mg / L rifampicin and 100 mg / L kanamycin, and mix 1 volume part of Agrobacterium containing target gene pBI121 vector with 500 volume parts of YEP culture medium , In a constant temperature shaker at 160 revolutions / min and 28°C for 24 hours, take 1 volume part of activated Agrobacterium containing the target gene pBI121 vector and 100 volume parts of YEP culture medium and mix it at 160 revolutions / min. Cultivate for 16 hours on a constant temperature shaker at 28°C for 16 hours until the OD600 value is 2.0, and collect Agrobacterium by...

Embodiment 3

[0041] To optimize the transgenic efficiency of Arabidopsis thaliana, the inflorescences on the Arabidopsis thaliana plants cultivated for 4 weeks were soaked with the transformation medium, and each Arabidopsis thaliana plant was wrapped with plastic wrap, the plants were cultivated, and the seeds were collected.

[0042] Among them, the specific configuration method of the transformation medium:

[0043] S1. Prepare YEP culture medium containing 50 mg / L rifampicin and 100 mg / L kanamycin, and mix 1 volume part of Agrobacterium containing target gene pBI121 vector with 500 volume parts of YEP culture medium , Activate and cultivate for 20 hours in a constant temperature shaker at 28°C at 160 revolutions / min. Take 1 volume part of the activated Agrobacterium containing the target gene pBI121 vector and mix with 100 volume parts of YEP medium culture solution at 160 revolutions / min. Cultivate for 14 hours on a constant temperature shaker at 28°C for 14 hours until the OD600 value is ...

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Abstract

The invention discloses a method for optimizing the transgenosis efficiency of arabidopsis thaliana. The method comprises the following steps: preparing a transformation medium for impregnating an inflorescence on an arabidopsis thaliana plant; covering each arabidopsis thaliana plant with a fresh-keeping film; culturing the arabidopsis thaliana plant for 24h under a dark condition and then culturing the arabidopsis thaliana plant under the conditions that the light period is 24h, the humidity is 60 to 70 percent and the temperature is 22 DEG C; collecting seeds, wherein the light period is 24h, and the arabidopsis thaliana plant is illuminated for 16h and is cultured in a dark place for 8h. The method disclosed by the invention has the beneficial effects of repeatedly improving the utilization rate of the arabidopsis thaliana plants and the transformation medium and improving the transformation rate of the arabidopsis thaliana plants.

Description

Technical field [0001] The invention relates to the technical field of Arabidopsis transgene. More specifically, the present invention relates to a method for optimizing transgene efficiency in Arabidopsis. Background technique [0002] Arabidopsis thaliana is a small flowering plant widely distributed in Asia, Europe and North Africa. As the most widely used model plant in recent years, Arabidopsis thaliana has played an important role in the research of molecular genetics, botany and agricultural science. It is called the fruit fly in plants and is currently one of the five recognized model organisms. . [0003] The method commonly used to transform Arabidopsis with Agrobacterium is the flower dip method. In general, the Arabidopsis plants used for transformation need to be cultured under normal conditions for 4 weeks, select the first batch of Arabidopsis thaliana inflorescences that bolt and bloom, cultivate the Agrobacterium with the target gene, prepare the transformation ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/82A01H5/00A01H6/20
CPCC12N15/8205
Inventor 黄浩白隆华翟勇进韦莹
Owner GUANGXI BOTANICAL GARDEN OF MEDICINAL PLANTS
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