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Method for constructing efficient genetic transformation system of lilium lancifolium

A genetic transformation system, lily technology, applied in the field of constructing an efficient genetic transformation system for lily lily, can solve the problems of easy-to-infect bacteria, easy to produce chimeras, low transformation efficiency, etc. high efficiency effect

Active Publication Date: 2019-09-03
YANGTZE NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] Aiming at the above-mentioned deficiencies existing in the prior art, the object of the present invention is to provide a method for constructing a highly efficient genetic transformation system for lily lily, so as to solve the problems of low transformation efficiency, easy-to-contaminate bacteria, and easy generation of mosaic Combination problems, especially to provide technical support for the construction of efficient genetic transformation systems for lilies that are sensitive to Agrobacterium in part of the callus

Method used

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  • Method for constructing efficient genetic transformation system of lilium lancifolium
  • Method for constructing efficient genetic transformation system of lilium lancifolium
  • Method for constructing efficient genetic transformation system of lilium lancifolium

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Embodiment 1

[0056] 1. Activation of Agrobacterium liquid

[0057] Take the EHA105 (containing pLGNe plasmid) engineered bacterial strain stored at -80°C, and inoculate it in a 50mg L -1 and Rif40mg·L -1 YEB solid medium; after culturing in a constant temperature incubator at 28°C for 36-48h, pick a single colony strain with a pipette gun, inoculate it in YEB liquid medium, and shake it at 28°C at 180-250rpm for 24h. Then add it to YEB liquid medium at a volume ratio of 1:100 for activation, continue to shake and cultivate for about 12 hours, until the concentration of Agrobacterium bacteria is OD 600 The value is between 0.6-0.8. Then divide the bacterial solution into a sterile 50ml centrifuge tube, seal it with a parafilm, centrifuge at 5000rpm for 10 minutes, pour off the supernatant, and resuspend and dilute the bacterial cell EHA105 to OD with the improved resuspension solution 600 = 0.2.

[0058] 2. Obtaining of explants

[0059]Get the middle-layer small scales of the sterile ...

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Abstract

The invention discloses a method for constructing an efficient genetic transformation system of lilium lancifolium, which comprises the following steps: obtaining explants, activating Agrobacterium, infecting the Agrobacterium and co-cultivating Agrobacterium, and inducing bud differentiation and rooting. The method adopts a 2-3mm thin slice of the sterile small scale of lilium lancifolium as a transformation material, and establishes the efficient genetic transformation system of lilium lancifolium by medium culture improvement and optimization of Agrobacterium liquid concentration and infection time. The instantaneous transformation efficiency of lilium lancifolium is more than 80%, the stable conversion rate is above 25.00%, the conversion efficiency is high, the bacteria are not easilystained, and the chimera is not easily produced. The method solves the problem that the sterile small scale is used as the explant or a tTCL technology is used alone, and the genetic conversion efficiency of the lilium lancifolium is low; at the same time, the method overcomes the defect that lilium lancifolium callus is used as a transforming receptor, and the callus is too sensitive to Agrobacterium to finally obtain the transgenic plants, and the method has important guiding significance and reference value for the construction of the genetic transformation system of Agrobacterium-sensitive lilium lancifolium.

Description

technical field [0001] The invention belongs to the technical field of lily genetics and breeding, and in particular relates to a method for constructing an efficient genetic transformation system for lily lily. Background technique [0002] Lilium lancifolium Thunb. belongs to the Asiatic lily. Its plants are upright, with peculiar flower shape, orange-red color and black spots, so it is also called tiger skin lily and spotted lily. Lily lily is edible and medicinal. Its bulbs are white and fat, rich in protein, amino acid, starch, dietary fiber, alkaloids and minerals and other nutrients. Long-term consumption can lower blood sugar, clear lung and relieve cough, clear heart and calm the nerves and other effects, so it is processed into lily powder, dried lily, canned lily, etc. for sale in the market. Lily lily grows vigorously, has strong adaptability, and is resistant to viruses and fusarium. It can not only be used in landscaping, but also a good material for potted pl...

Claims

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Application Information

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IPC IPC(8): C12N15/82A01H5/00A01H6/56
CPCC12N15/8205C12N15/8261
Inventor 符勇耀杨利平李宏群刘建玲彭慎敏
Owner YANGTZE NORMAL UNIVERSITY
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