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Plant transgenic selection system using betaine aldehyde dehydrogenase (BADH) as marker gene and use thereof

A technology of marker gene and selection system, applied in the field of genetic engineering, can solve the problems of high price, high cost, and limit the widespread use of genetic transformation methods, and achieve the effect of reducing cost input

Inactive Publication Date: 2012-01-04
BEIJING UNIV OF AGRI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although this genetic transformation technology has been proven to be feasible, the transformation method uses betaine aldehyde to provide selection pressure, and betaine aldehyde is an extremely expensive selection agent, and the international market price is as high as 1,000 US dollars per gram The domestic price also reaches 2500-3000 yuan / gram, resulting in extremely high cost of this kind of genetic transformation, which in turn limits the widespread use of this genetic transformation method

Method used

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  • Plant transgenic selection system using betaine aldehyde dehydrogenase (BADH) as marker gene and use thereof
  • Plant transgenic selection system using betaine aldehyde dehydrogenase (BADH) as marker gene and use thereof
  • Plant transgenic selection system using betaine aldehyde dehydrogenase (BADH) as marker gene and use thereof

Examples

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Effect test

Embodiment 1

[0048] Example 1: Using BADH gene as a marker gene and using NaCl as a selection agent to carry out genetic transformation process of Prunus bicolor

[0049] 1. Determining the NaCl concentration that inhibits the differentiation of adventitious buds at the cotyledon nodes of E. bicolor: the explants adopted according to the genetic transformation process of E. bicolor are cotyledon nodes, and the cotyledon nodes are at MS+6-BA 0.2mg / L+3% sucrose +6g / L agar medium has the best effect on adventitious bud differentiation. Based on this medium, configure medium containing different concentrations of NaCl, such as: MS+6-BA0.2mg / L+3% sucrose+6g / L agar+NaCl 0.4g / L, MS+6-BA 0.2mg / L+3% sucrose+6g / L agar+NaCl0.6g / L, MS+6-BA 0.2mg / L+3% sucrose+6g / L agar+NaCl 0.8g / L, MS+6-BA 0.2mg / L+3% sucrose+6g / L agar+NaCl 1.0g / L and other series of media, observe the adventitious bud differentiation of cotyledon nodes on these media In this case, since the cotyledon nodes of the two-colored ramif...

Embodiment 2

[0053] Example 2: Genetic transformation of tobacco with BADH gene as marker gene and NaCl as selection agent:

[0054] 1. Understand the effect of NaCl concentration on the differentiation ability of adventitious buds of tobacco leaves: the suitable medium for the differentiation of tobacco leaves is 1 / 2MS+6-BA 2.0mg / L+NAA 0.1mg / L+3% sucrose+6g / L agar. On the basis of this medium, add 0, 0.4, 0.8, 1.2, 1.6, 2.0, 3.0g / L NaCl respectively, observe the growth performance of tobacco leaves on various media, and determine to add 2g / L NaCl to the medium. L NaCl can inhibit the differentiation of adventitious buds in tobacco leaves;

[0055] Add 2g·L to the appropriate medium for tobacco leaf differentiation 1 / 2MS+6-BA 2.0mg / L+NAA 0.1mg / L+3% sucrose+6g / L agar -1 NaCl, to obtain the selection medium, the tobacco leaves after the transformation of the exogenous gene with BADH as the marker gene are placed in the selection medium for screening;

[0056] Figure 10 For the differen...

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Abstract

The invention discloses a plant transgenic selection system using betaine aldehyde dehydrogenase (BADH) as a marker gene and use thereof and belongs to the technical field of gene engineering. In the system, BADH is used as a marker gene of an exogenous gene and NaCl is used as a selecting agent. The use comprises the following steps: (1) determining NaCl concentration which can just inhibit plant tissue differentiation; (2) transferring the exogenous gene which contains the BADH marker gene into a plant cell or tissue; and (3) culturing the transgenic tissue or cell into which the BADH marker gene is transferred on a culture medium in which the NaCl concentration determined by the step (1), obtaining the transgenic tissue or cell into which the BADH marker gene is transferred into if the transgenic tissue or cell can grow, and confirming by molecular detection. The invention has the advantages of greatly lowering the cost investment in a transgenic test, solving the safety problem of transgenic organisms, and providing low cost selection for the construction of a safe and effective selection system for plant genetic transformation.

Description

Technical field: [0001] The invention belongs to the technical field of genetic engineering, and in particular relates to a plant transgenic selection system using BADH as a marker gene and its application. Background technique: [0002] The use of genetic engineering technology to achieve genetic improvement of plant traits has become a new trend in the cultivation of new plant varieties. In the process of plant transgenics, the adoption of an effective selection system is a key link in the success of plant transgenics. The most commonly used selection systems are the herbicide selection system and the antibiotic selection system. Although these two selection systems have been successfully used in the genetic transformation of a variety of plants, the adoption of these two selection systems has raised public concerns about the safety of GMOs. An important cause of sexual concern. On the one hand, there is concern that antibiotic resistance genes in transgenic plants will ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/09A01H5/00
Inventor 杨晓红刘雯陈晓阳解小娟冷平生
Owner BEIJING UNIV OF AGRI
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