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Cymbidium goeringii CgWRKY65 gene and application thereof

A gene and amino acid technology, applied to the Chunlan CgWRKY65 gene and its application fields, can solve the problems of reduced activity, chromosome influence, gene inactivation, etc., and achieve the effect of prolonging the vegetative growth cycle and being widely used.

Active Publication Date: 2020-07-17
NANJING FORESTRY UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

When the foreign gene enters the chromosome, the foreign protein encoded by the gene is produced, and the foreign protein or enzyme leads to the enhancement or reduction of the activity of other enzymes or proteins through a series of reactions. In addition, when the foreign gene is embedded in a certain part of the chromosome When a segment is formed, it will definitely affect the entire chromosome, thereby affecting the activity or regulatory mechanism of other genes.
[0005] Even if the same gene is inserted, due to different insertion sites or different chromosome groups, it will inevitably affect the chromosome where it is located, and further lead to an impact on the gene on the chromosome where it is located, and slight changes in the gene may affect its regulatory mechanism , lead to changes such as gene inactivation, activity reduction, activity enhancement, etc., and also cause differences in the role of genes, and the role of genes can only be obtained through verification, not through inference

Method used

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  • Cymbidium goeringii CgWRKY65 gene and application thereof
  • Cymbidium goeringii CgWRKY65 gene and application thereof
  • Cymbidium goeringii CgWRKY65 gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] The material used in this example is the leaves of Chunlan 'Songmei', which are quickly frozen in liquid nitrogen after harvest and stored in an ultra-low temperature refrigerator (-80°C).

[0024] 1) Extraction of total RNA from Chunlan leaves

[0025] According to the instructions of the TaKaRa plant total RNA extraction kit, the specific operation is as follows:

[0026] The ultra-low temperature frozen leaves of Chunlan were quickly transferred to a mortar pre-cooled with liquid nitrogen, and the tissue was ground with a pestle, during which liquid nitrogen was continuously added until it was ground into powder; the powdered sample was added to a 450 μl In a 1.5 mL sterilized tube of Buffer PE, pipette repeatedly until there is no obvious precipitation in the lysate; centrifuge the lysate at 12,000 rpm at 4°C for 5 min; carefully pipette the supernatant into a new 1.5 mL sterilized tube . Add 1 / 10 volume of Buffer NB to the supernatant, shake and mix with Vortex, ...

Embodiment 2

[0048] Research indicates WRKY65 The gene was expressed in every tissue and organ of Chunlan ( figure 2 a), but the gene has the highest expression level in spring orchid, indicating that the gene is functionally active in flowers. Through the expression analysis of the leaves of Chunlan ABA stress treatment ( figure 2 b), to prove CgWRKY65 Genes play an important regulatory role in the ABA stress response of Chunlan leaves.

[0049] The plant material used in this embodiment is Arabidopsis ( Arabidopsis thaliana ) Col (Columbia) wild-type seeds.

[0050] The Escherichia coli strain used in this example was Trans5α; the Agrobacterium strain was GV3101, which were used to transform Arabidopsis thaliana; the plant expression vector used in the experiment was pBI121. The strains used were purchased from Quanshijin Biotechnology Company and Price Biotechnology Company.

[0051] 1) CgWRKY65 Construction of gene overexpression vector

[0052] With embodiment 1 obtaining ...

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Abstract

The invention discloses a cymbidium goeringii CgWRKY65 coding gene and application thereof. The nucleotide sequence of the CgWRKY65 gene is shown as SEQ ID NO.1, and the amino acid sequence of an expression protein is shown as SEQ ID NO.2. Through cloning and identification, gene expression analysis and functional verification of the CgWRKY65 gene of a cultivated cymbidium goeringii variety Songmei, the invention discovers that: in a seedling stage of CgWRKY65 transgenic Arabidopsis thaliana, compared with wild Arabidopsis thaliana WT, the whole CgWRKY65 transgenic plant is short, has yellowish leaves, and has a tendency of premature senescence, so that the CgWRKY65 gene may have wide application in orchid nutrition and reproduction improvement.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, in particular to a spring orchid CgWRKY65 Genes and their applications. Background technique [0002] Orchidaceae (Orchidaceae) is one of the largest families of flowering plants, with more than 25,000 species in the world, accounting for about 10% of all flowering plants. Chunlan ( Cymbidium goeringii ) belongs to the small-flowered ground orchid species in the Orchidaceae genus. It has a peculiar flower shape, elegant flower color, quiet flower fragrance, beautiful leaf appearance, and high ornamental and economic value. Chunlan has high requirements on the growth environment. During the growth process, it is easily affected by harsh environments such as high temperature, low temperature, and drought. Therefore, it is of great significance to study the molecular mechanism of plant response to abiotic stress and identify the genes with stress resistance function for Chunlan...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/82C07K14/415A01H5/12A01H6/20
CPCC07K14/415C12N15/8261
Inventor 胡凤荣刘倩王连平徐子涵黎猛
Owner NANJING FORESTRY UNIV
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