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Method for silencing TRV-based virus induced primula malacoides genus plant gene

A technology of gene silencing and primrose, applied in the field of plant genetic engineering, can solve problems such as the research on the function of genes limiting the S site, and achieve the effects of short research period, high phenotype efficiency and simple method

Active Publication Date: 2019-11-26
BEIJING FORESTRY UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Since the genetic transformation system of Primula miltiorrhiza has not been established, the research on the function of the gene at the S site is limited.

Method used

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  • Method for silencing TRV-based virus induced primula malacoides genus plant gene
  • Method for silencing TRV-based virus induced primula malacoides genus plant gene
  • Method for silencing TRV-based virus induced primula malacoides genus plant gene

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Embodiment 1

[0056] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the present invention. Unless otherwise specified, the examples are all in accordance with conventional experimental conditions, such as Sambrook et al. Molecular Cloning Experiment Manual (Sambrook J & Russell DW, Molecular Cloning: a Laboratory Manual, 2001), or in accordance with the conditions suggested by the manufacturer's instructions. Example 1 TRV-based virus-induced method for gene silencing of Primula miltiorrhiza

[0057] 1. Acquisition of experimental materials: Seeds are sown on MS medium, and the seedlings are cultivated to 6-9 true leaves before being bottled, and the seedlings are cultivated in an artificial climate chamber after being out of the bottle for subsequent use. Culture conditions: light 300μmol·m -2 ·s -1 , 16 hours of light, 8 hours of darkness, daytime temperature of 22°C, darkness of 18°C, and humidity of 60%.

[0058] 2. Rec...

Embodiment 2

[0082] Example 2 Using VIGS to analyze the function of the auxin export protein gene in Primula miltiorrhiza

[0083] Auxin is involved in regulating various processes in plant growth and development, such as cell division and cell elongation. Physiological pleiotropy was exhibited at the tissue, organ and whole plant levels. The transport of auxin between cells depends on the carrier located in the cell membrane. Auxin export protein PIN-FORMED (PIN) is an important class of proteins that transport auxin to the extracellular transmembrane. It has been reported that silencing the TaPIN1 gene in wheat leads to dwarf plants (Kanwardeep, 2011), while the PtPIN8 gene is negatively correlated with the root hair length of Citrus citrus (Liu et al., 2108). The PIN gene may affect the elongation or division of cells by affecting the transport and redistribution of auxin, and ultimately affect the height or length of the whole plant or organ. In the early stage of the present invent...

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Abstract

The invention provides a method for silencing a TRV-based virus induced primula malacoides genus plant gene. The method comprises the steps that A, according to a primula malacoides genus plant targetgene, a primer is designed, and a DNA fragment of the target gene is amplified; B, the DNA fragment is connected to a pTRV2 vector, and a recombinant vector is obtained; and C, a pTRV1 and the recombinant vector transform agrobacterium correspondingly, and obtained transformants commonly infect plants. According to the method, it is firstly proved that TRV can be successfully applied to gene silencing and functional identification of primula forbesii, and a foundation is laid for gene functional identification of primula malacoides genus plants. The method is simple, the research period is short, a full-length gene or a transformation system is not needed, the efficiency for obtaining a target phenotype is high, and quick functional identification of a large amount of genes can be achieved. compared with an existing VIGS system, the lasting time of phenotype of the method is long, phenotype changes appear on different parts of a plant, and the method has important significance on genetic research of self-incompatibility of primula malacoides genus heterotype.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, and in particular relates to a method for gene silencing of Primula plants induced by TRV-based viruses. Background technique [0002] Gene silencing is an important means of regulating gene expression in eukaryotic cells. Virus-induced gene silencing (VIGS) is a type of post-transcriptional gene silencing (PTGS). Due to the introduction of exogenous genes, the endogenous plants in plants that are consistent or highly homologous to the exogenous gene Specific degradation of mRNA sequences. Compared with transgenic technology, VIGS technology has the advantages of simple operation, fast and effective, and is a better reverse genetics tool for gene function verification. [0003] The artificially improved tobacco rattle virus (TRV) is currently the most widely used VIGS vector. Compared with other viral vectors, TRV vectors can effectively transmit silencing signals to the grow...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/82C12N15/66A01H5/00A01H6/00
CPCC12N9/001C12N15/66C12N15/8218C12Y103/99028
Inventor 潘会堂司未佳张启翔申建双刘颖程堂仁王佳
Owner BEIJING FORESTRY UNIVERSITY
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