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Agrobacterium tumefaciens-mediated efficient genetic transformation system for lilium tenuifolium scales

A genetic transformation system and technology mediated by Agrobacterium, applied in the field of establishing an efficient genetic transformation system of lily lily mediated by Agrobacterium, can solve the problems of poor genetic stability of transformed plant traits, unstable T-DNA integration, and insensitivity to Agrobacterium Sex and other issues, to achieve the effect of strong direct regeneration, low false positive rate, and avoidance of mutation

Inactive Publication Date: 2020-05-26
NORTHEAST FORESTRY UNIVERSITY
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  • Abstract
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  • Claims
  • Application Information

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Problems solved by technology

However, lily is a monocotyledonous plant, and some problems may arise when using Agrobacterium-mediated gene transformation, such as insensitivity to Agrobacterium, unstable T-DNA integration, poor genetic stability of transformed plant traits, etc.

Method used

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  • Agrobacterium tumefaciens-mediated efficient genetic transformation system for lilium tenuifolium scales
  • Agrobacterium tumefaciens-mediated efficient genetic transformation system for lilium tenuifolium scales
  • Agrobacterium tumefaciens-mediated efficient genetic transformation system for lilium tenuifolium scales

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Embodiment Construction

[0034] The construction of the Agrobacterium-mediated genetic transformation system of Lilium tenuifolia scales is further illustrated by the following examples. The Agrobacterium tumefaciens strain used in the examples is EHA105, which contains the plant expression vector PBI121, and carries the green fluorescent protein gene GFP.

[0035] 1. Antibiotic sensitivity test

[0036] Will be cut into 0.5cm 2 Small and small scales of Lily lily are inoculated on medium (MS+1.5mg / L 6-BA+0.5mg / L NAA+400mg / L Cef) supplemented with different concentrations of Kan (0, 100, 110, 120, 130 mg / L) ) For culturing, regularly observe the growth status of the small scales and count the browning rate to determine the optimal screening concentration of Kan for small scales.

[0037] Table 1 Kan concentration screening

[0038]

[0039] It can be seen from Table 1 that kan has an inhibitory effect on the differentiation of small scales. In the medium without kan, the browning rate of the small scales i...

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Abstract

The invention discloses an agrobacterium tumefaciens-mediated efficient genetic transformation system for lilium tenuifolium scales. The system comprises the following steps: screening proper concentration of antibiotic kanamycin; preparing a bacterial solution; carrying out pre-culture; performing infection; carrying out co-culture; screening and culturing; obtaining resistant seedlings; pCR detection of transgenic plants. According to the invention, sterile flakes of lilium tenuifolium are used as a transformation material, main transformation conditions in the agrobacterium-mediated transformation process are optimized, an agrobacterium-mediated efficient genetic transformation system of lilium tenuifolium flakes is established, a green fluorescent protein gene GFP is transferred into lilium tenuifolium, and the transformation rate reaches 16.13%. The method has the advantages that the lilium tenuifolium scales are used as a direct regeneration path of a transformation receptor, theregeneration rate is high, the mutation rate and the false positive rate are low, the transformation period is short, complete plants can be obtained in about 3.5 months, the transformation rate is high, and a foundation is laid for molecular breeding, germplasm resource creation and variety improvement of lilies.

Description

Technical field [0001] The invention relates to a method for genetic transformation of lily in the field of biotechnology, in particular to a method for establishing an Agrobacterium-mediated high-efficiency genetic transformation system for lily lily. Background technique [0002] Lilium pumilum (Lilium pumilum), also known as Shandan, is native to the cold regions of northern China. The petals are rolled outwards, the flower color is bright red, and the ornamental value is extremely high. It can be directly used in gardens and landscapes; and has strong drought and resistance The ability to resist cold and salinity is an important parent of lily resistance breeding. In recent years, the research on Lilium tenuifolia has mainly focused on hybrid breeding and taxonomy, but there are few reports on its genetic engineering. Using conventional breeding techniques to improve plant varieties is not only time-consuming, laborious, and difficult, but the use of genetic engineering meth...

Claims

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Application Information

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IPC IPC(8): C12N15/82C12N15/65A01H4/00A01H5/00A01H6/56
CPCC12N15/8205C12N15/65A01H4/00
Inventor 张彦妮刘彬王营
Owner NORTHEAST FORESTRY UNIVERSITY
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