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Tomato ascorbic acid biosynthetic gene PMI and application thereof

A technology of ascorbic acid and biosynthesis, applied in the field of genetic engineering, can solve the problems of lack of PMIs genetic test evidence and few reports on regulatory mechanisms

Pending Publication Date: 2022-08-05
武汉楚为生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0011] (2) At present, there is a lack of specific experimental evidence on how PMIs genes play a role in the regulation of tomato ascorbic acid synthesis and metabolism
At present, there are few reports on the regulatory mechanism of genes related to ascorbic acid biosynthesis

Method used

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  • Tomato ascorbic acid biosynthetic gene PMI and application thereof
  • Tomato ascorbic acid biosynthetic gene PMI and application thereof
  • Tomato ascorbic acid biosynthetic gene PMI and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0083] 1) Plant material:

[0084] The tomato material is (Solanum lycopersicum) conventional line AC (preserved in this laboratory), which is used for gene cloning and genetic transformation. AC was potted in spring. After the fruit set matured, the roots, stems, leaves, flowers and fruit samples of different developmental stages were collected and quick-frozen in liquid nitrogen, and stored in a -70°C ultra-low temperature freezer for analysis of tissue expression patterns of PMIs.

[0085] 2) PMIs tissue expression profile and analysis of ascorbic acid content in each tissue:

[0086] Primer qPCR primers for PMI gene were designed using Primer5 software. A 10 μL reaction system was used, with EF1a as the internal reference (Lovdal and Lillo, 2009), and the Roche fluorescence quantitative PCR instrument LC480 was used for determination and analysis. The system includes Premix Ex Taq TM (2×) 5 μL, 0.5 μL of forward and reverse primers, 4 μL of template. Each sample was r...

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Abstract

The invention belongs to the technical field of gene engineering, and discloses a tomato ascorbic acid biosynthetic gene PMI and application, the tomato ascorbic acid biosynthetic gene PMI is PMI1 and PMI2, and DNA of ORF of PMI1 and PMI2 is SEQ ID NO: 1 and SEQ ID NO: 2; proteins coded by the tomato ascorbic acid biosynthetic gene PMI comprise SEQ ID NO: 3 and SEQ ID NO: 4. According to the invention, a tomato conventional strain AC is taken as an object, an ascorbic acid synthesis Smirnoff pathway key gene is cloned, a corresponding genetic transformation vector is constructed, and the vector is transferred into the tomato strain AC by using an agrobacterium tumefaciens-mediated genetic transformation method, so that the expression of a target gene is excessive or inhibited; the ascorbic acid content of the obtained transgenic tomato is analyzed, and the effect of the genes in tomato ascorbic acid content regulation and control is evaluated.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, in particular to a tomato ascorbic acid biosynthesis gene PMI and its application. Background technique [0002] Currently, the closest existing technology: [0003] Ascorbic acid (Ascorbic Acid, AsA), also known as vitamin C (Vc), has a very important role in the body, such as antioxidant, cofactor of important enzymes and so on. Studies in plants have found that AsA is positively correlated with plant stress resistance. Increasing the content of endogenous AsA can improve plant stress resistance, such as cold tolerance. At the same time, ascorbic acid is an important substance for human and animal to maintain growth and development. However, primates (such as humans) lack the last enzyme in the AsA biosynthetic pathway, so they cannot synthesize AsA, and can only obtain sufficient amounts of AsA from plants (Jain et al., 2000), and the fresh fruit is ascorbic acid main source. ...

Claims

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Application Information

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IPC IPC(8): C12N15/61C12N9/90C12N15/84C12N15/66A01H5/08A01H6/82C12Q1/6851C12Q1/6895
CPCC12N9/90C12N15/8205C12N15/8243C12Q1/6851C12Q1/6895C12Y503/01008C12Q2600/13C12Q2600/158C12Q2531/113C12Q2545/101C12Q2563/107
Inventor 张余洋叶志彪郑伟
Owner 武汉楚为生物科技有限公司
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