Method for quickly obtaining transgenic eggplant plants

A technology of transgenic and eggplant, applied in botany equipment and methods, horticultural methods, genetic engineering, etc., can solve the problems of low genetic transformation efficiency of eggplant, achieve the effect of solving unstable quality of regenerated buds, shortening the transformation cycle, and quickly obtaining

Inactive Publication Date: 2019-09-27
HARBIN CITY ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve the problem of low genetic transformation efficiency of eggplant, the present invention provides a method for rapidly obtaining transgenic plants of eggplant

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] The present embodiment provides a kind of method that obtains eggplant transgenic plant rapidly, and the steps are as follows:

[0027] Step 1. Pre-cultivate eggplant explants on the regeneration medium, which contains 0.1-0.5 μg / mL thiadidiazuron and 10-15 μg / mLN 6 -[isoamyl]-adenine;

[0028] Step 2, Agrobacterium activation: prepare a strain of Agrobacterium carrying recombinant DNA, its recombinant DNA is cryIIIA gene and selectable marker gene, the marker gene has kanamycin resistance to the cells transformed by recombinant DNA, this strain Agrobacterium is inoculated on YEB solid medium containing kanamycin, cultured in dark at a certain temperature until a single colony of Agrobacterium grows, and single colony is inoculated in YEB liquid medium containing kanamycin, at a certain temperature Carry out shaking culture, transfer the bacterial liquid obtained from the shaking culture into a centrifuge tube, centrifuge at 4000r / min, collect the bacteria and suspend ...

Embodiment 2

[0033] The present embodiment provides a kind of method that obtains eggplant transgenic plant rapidly, and the steps are as follows:

[0034] Step 1. Pre-cultivate eggplant explants on the regeneration medium, which contains 0.1-0.5 μg / mL thiadidiazuron and 10-15 μg / mLN 6 -[isoamyl]-adenine;

[0035] Step 2, Agrobacterium activation: prepare a strain of Agrobacterium carrying recombinant DNA, its recombinant DNA is cryIIIA gene and selectable marker gene, the marker gene has kanamycin resistance to the cells transformed by recombinant DNA, this strain Agrobacterium is inoculated on YEB solid medium containing kanamycin, cultured in dark at a certain temperature until a single colony of Agrobacterium grows, and single colony is inoculated in YEB liquid medium containing kanamycin, at a certain temperature Carry out shaking culture, transfer the bacterial liquid obtained from the shaking culture into a centrifuge tube, centrifuge at 4000r / min, collect the bacteria and suspend ...

Embodiment 3

[0041] The present embodiment provides a kind of method that obtains eggplant transgenic plant rapidly, and the steps are as follows:

[0042] Step 1. Pre-cultivate eggplant leaf explants on the regeneration medium, the components of the regeneration medium include: 0.1-0.5 μg / mL thiadidiazuron, 10-15 μg / mLN 6 -[Isoamyl]-Adenine, 15~20mM NH 4 NO 3 , 12~16mM KNO 3 , 2~2.5mM CaCl 2 2H 2 O, 1 ~ 1.2mM MgSO 4 ·7H 2 O, 0.8 ~ 1.0mM KH 2 PO 4 , 3~4μM KI, 0.05~0.08mMH 3 BO 3 , 0.05~0.08mM MnSO 4 4H 2 O, 20-25μM ZnSO 4 ·7H 2 O, 0.5~0.8μM Na 2 MoO 4 2H 2 O, 0.05~0.08μM CuSO 4 ·5H 2 O, 0.05~0.08μM CoCl 2 ·6H 2 O, 0.05-0.08mM EDTA, pH5.7-5.9.

[0043] Step 2, Agrobacterium activation: prepare a strain of Agrobacterium carrying recombinant DNA, its recombinant DNA is cryIIIA gene and selectable marker gene, the marker gene has kanamycin resistance to the cells transformed by recombinant DNA, this strain Agrobacterium is inoculated on YEB solid medium containing kanamyc...

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Abstract

The invention relates to a method for quickly obtaining transgenic eggplant plants and belongs to the technical field of a transgenic eggplant plant obtaining method. In order to solve the problem of low eggplant genetic transformation efficiency, the invention provides the method for quickly obtaining the transgenic eggplant plants, which comprises the following steps: firstly, pre-culturing eggplant explants on a regenerative culture medium, activating agrobacterium tumefaciens carrying cryIIIA genes, infecting the eggplant explants with the agrobacterium tumefaciens and culturing the infected eggplant explants on the regenerative culture medium to transform explant cells with the agrobacterium tumefaciens; then transferring the explants onto a selective culture medium for further culture so as to form adventitious buds and further form the complete plants. The method provided by the invention has the benefits that both the regenerative culture medium and the selective medium contain thiabendron and N6-[isoamene]-adenine, so that the induction efficiency is high, the quantity of the adventitious buds formed by the explants can be increased, and the regeneration efficiency growth rate can reach 163 percent; according to the method provided by the invention, the transformation efficiency of transgenic eggplant breeding is improved, and the transformation period is shortened.

Description

technical field [0001] The invention belongs to the technical field of methods for obtaining transgenic plants, and in particular relates to a method for rapidly obtaining transgenic plants of eggplant. Background technique [0002] Eggplant is rich in nutrients and contains protein, fat, carbohydrates, vitamins, calcium, phosphorus, iron and other nutrients. It is one of the common vegetables on the table. Eggplant is vulnerable to various diseases such as Verticillium wilt, bacterial wilt, and cotton blight, as well as nematodes and various insects during the planting process, resulting in large-scale production reduction and quality decline of eggplant. Although a few wild germplasm resources are disease-resistant materials, their natural resistance is not enough to completely resist pests and diseases, and they cannot be fully utilized in the breeding process. [0003] In traditional eggplant breeding, due to the existence of linked toxic genes, resulting in incompatibl...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/82A01H4/00A01H5/00A01H6/82
CPCC12N15/8205C12N15/8286A01H4/001A01H4/008
Inventor 李烨曹必好姜景彬许向阳赵婷婷谢立峰陈柏杰
Owner HARBIN CITY ACAD OF AGRI SCI
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