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Capsella bursa-pastoris peroxidase gene and application of capsella bursa-pastoris peroxidase gene to improvement of cold resistance of economic plants

A shepherd's purse peroxidase and enzyme gene technology, which is applied in the field of plant molecular biology and can solve the problem of not finding the shepherd's purse peroxidase gene and the like

Inactive Publication Date: 2016-10-12
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

There is no report on the use of shepherd's purse peroxidase gene in cultivating cold-resistant plants

Method used

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  • Capsella bursa-pastoris peroxidase gene and application of capsella bursa-pastoris peroxidase gene to improvement of cold resistance of economic plants
  • Capsella bursa-pastoris peroxidase gene and application of capsella bursa-pastoris peroxidase gene to improvement of cold resistance of economic plants
  • Capsella bursa-pastoris peroxidase gene and application of capsella bursa-pastoris peroxidase gene to improvement of cold resistance of economic plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Example 1 Isolation and identification of shepherd's purse peroxidase gene

[0069] 1. Cultivation of shepherd's purse seedlings

[0070] The shepherd's purse seeds come from Shanghai Seed Company. After the shepherd's purse seeds are sterilized with 75% ethanol, they are sown in the 0 On the culture medium, place the seedlings at 26°C for 2w and then transplant the seedlings into the soil (vermiculite: black soil: perlite = 9:3:1) to continue to grow. During the growth process, the photoperiod is 16h light, 8h dark, humidity Keep it constant and moderate.

[0071] 2. Extraction of Genomic DNA and RNA from Shepherd's Purse Leaf

[0072] The leaves of shepherd's purse of appropriate size were ground into powder in liquid nitrogen, and the total RNA and whole genome DNA of shepherd's purse were extracted by plant RNA extraction kit (CW0588) (Beijing Kangwei Century Biotechnology Co., Ltd.) and CTAB method, respectively. Agarose gel electrophoresis was used to detect ...

Embodiment 2

[0080] Example 2 Bioinformatics analysis of shepherd's purse peroxidase gene

[0081] 1. Gene structure analysis

[0082] Comparing the obtained sequence with the CDS sequence of shepherd's purse peroxidase gene (AY566573), it was found that the genome sequence of shepherd's purse contains 3 introns, and all 3 introns contain the gt-ag intron conserved sequence. Intron, the CDS sequence of shepherd's purse peroxidase gene contains 9811bp.

[0083] 2. Homology analysis

[0084] Use http: / / www.ncbi.nlm.nih.gov / blast to search by Protein-Protein BLAST and other peroxidase protein family members including Arabidopsis ( A. thaliana ), peroxidase (NM100405and NP188814), soybean ( Glycine max ) in peroxidase (AAD11481), Eucommia ( Eucommiaulmoides ), peroxidase (AAU04879) in spinach ( Spinaciaoleracea Peroxidase (CAA76374) in rice ( Oryza sativa ), peroxidase (CAH69331) in Ziping ( Spirodela polyrrhizae ), peroxidase (CAA80502) in tobacco ( Nicotiana tabacum Peroxidas...

Embodiment 3

[0087] The expression specificity analysis of embodiment 3 shepherd's purse peroxidase

[0088] 1. Various treatment conditions of shepherd's purse plants

[0089] (1) Cold induction treatment: move the shepherd's purse seedlings grown for 4w under long-day conditions at 22°C to 4°C for 4h, 8h, and 24h respectively;

[0090] (2) Cold acclimatization treatment: put shepherd's purse seedlings grown under long-day conditions at 22°C for 4w, and put them in 12°C for 4 days, 4°C for 4 days, and 0°C for 2 hours;

[0091](3) Hormone treatment: The shepherd's purse seedlings grown for 4w under long-day conditions at 22°C were soaked in 5 μM GA, 300 μM MeJA, 20 μM IAA, and 100 μM ABA aqueous solutions for 1 hour, 6 hours, and 24 hours, respectively;

[0092] (4) Ion solution treatment: use 80mM KCl and 50mM MgCl respectively to shepherd's purse grown for 4w under long-day conditions at 22°C 2 , 5mM ZnCl 2 , 30mM LiCl, 0.1mM CuCl 2 , 80mM CaCl 2 Soak in aqueous solution for 4h, 8h...

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Abstract

The invention belongs to the technical field of plant molecular biology, in particular to a capsella bursa-pastoris peroxidase gene and application of capsella bursa-pastoris peroxidase gene to improvement of cold resistance of economic plants. The nucleotide sequence of the capsella bursa-pastoris peroxidase functional gene is shown in SEQ ID NO: 1, and the capsella bursa-pastoris peroxidase gene is encoded by the nucleotide sequence, is expressed through low temperature induction, is under-expressed at normal temperature, and further promotes cells to generate related low-temperature induced genes through regulation of active oxygen in plant cells after expression, so as to finally improve the cold resistance of the plants. An over-expressed vector constructed by utilizing the gene segment can be over-expressed in the plants, and then cold resistant gene expression can be regulated and controlled, so as to protect the plant cells from being destroyed by low temperature. The invention further provides an application method for cultivating the cold resistant plants based on the gene, and the application method is used for improvement of varieties of crops with economic value.

Description

technical field [0001] The invention belongs to the technical field of plant molecular biology, and specifically relates to a shepherd's purse peroxidase gene, a preparation method thereof, a recombinant plasmid of the gene, and its application to plant adversity regulation, especially to the inheritance of economic plant cold resistance genes Transformation to increase plant cold resistance. Background technique [0002] Temperature is an indispensable environmental condition in the process of plant growth. Plants, as a kind of fixed growth organisms, cannot avoid adverse environments by walking and moving, and are more restricted by environmental factors. Low temperature freezing damage is a serious natural disaster in agricultural production. It will not only limit the planting range of crops, but also cause crop yield reduction, causing huge losses of crops every year. Therefore, the research on plant cold resistance should transform the genetic characteristics of crops ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53C12N9/08C12N15/82C12N15/84C12N5/10A01H5/00
CPCC12N9/0065C12N15/8273C12Y111/01
Inventor 林娟李伟伟陈虎魏东晖
Owner FUDAN UNIV
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