Plant stress tolerance-related protein GmNF-YC9 as well as encoding gene and application thereof

A technology of encoding genes and genes, which is applied in the direction of plant gene improvement, application, plant peptides, etc., and can solve the problems of comprehensive improvement of plant stress resistance that are difficult to achieve

Active Publication Date: 2014-01-15
INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] Since the stress tolerance of plants is a complex trait regulated by multiple genes, it is difficult to achieve comprehensive improvement of plant stress resistance by introducing a single functional protein gene

Method used

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  • Plant stress tolerance-related protein GmNF-YC9 as well as encoding gene and application thereof
  • Plant stress tolerance-related protein GmNF-YC9 as well as encoding gene and application thereof
  • Plant stress tolerance-related protein GmNF-YC9 as well as encoding gene and application thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Embodiment 1, the cloning of GmNF-YC9 gene

[0052] 1. Isolation of mRNA

[0053] Soybean Tiefeng 8 (Glycine max L.) four-leaf stage seedlings grown in hydroponics for 10 days were drought-treated for 2 hours, quickly frozen with liquid nitrogen, and stored at -80°C for later use. mRNA was isolated using Quikprep Micro mRNA Purification Kit (Pharmacia). First-strand cDNA synthesis was performed with Reverse Transcriptase XL (AMV). The ds cDNA was synthesized by SMART method, and the PCR products were detected by 1.0% agarose gel electrophoresis.

[0054] 2. Acquisition of the full-length sequence of the GmNF-YC9 gene

[0055] The full-length cDNA sequence of the nuclear transcription factor C family gene of soybean CCAAT-box was obtained by 5'RACE and 3'RACE methods, as shown in Sequence Listing Sequence 2, the name is GmNF-YC9 gene, and its open reading frame is the self-sequence listing sequence From the 34th to the 897th nucleotide of the 5' end of 2, the amino a...

Embodiment 2

[0056] Embodiment 2, real-time fluorescent quantitative PCR analysis of the expression characteristics of GmNF-YC9 gene

[0057] 1. Coercion treatment

[0058] Be the No. 8 seedling of soybean Tiefeng of 10 days with potted seedling age, carry out following processing:

[0059] (1) Abscisic acid treatment ( figure 1 A): Placed in 100 μM abscisic acid (ABA) solution, cultured under light for 0.5 hours, 1 hour, 2 hours, 5 hours, 12 hours, and 24 hours, then took them out and quick-frozen them with liquid nitrogen, and stored them at -80°C for later use.

[0060] (2) Drought treatment ( figure 1B): Take out and absorb the water on the root, place it on dry filter paper, and cultivate it in a drought for 0.5 hours, 1 hour, 2 hours, 5 hours, 12 hours, and 24 hours. After that, take out the material, freeze it with liquid nitrogen, and store it at -80°C for later use .

[0061] (3) Low temperature treatment ( figure 1 C): placed in a 4°C incubator, cultured under light for 0.5 ...

Embodiment 3

[0079] Embodiment 3, the activation characteristic of GmNF-YC9

[0080] The main principle of using the yeast one-hybrid system to demonstrate the activation properties of transcription factors is as follows: figure 2 As shown, the CCAAT cis-acting element and the mutant CCAAT cis-acting element were constructed to the upstream of the basic promoter Pmin (minimal promoter) of the pHISi-1 vector and the pLacZi vector, respectively, and the reporter genes (His3, LacZ and URA3). When the expression vector YEP-GAP (without activation function) connected with the target gene encoding the transcription factor is transformed into the yeast cells connected with the CCAAT cis-acting element and the mutant CCAAT cis-acting element, if the mutant CCAAT The reporter gene in yeast cells with cis-acting elements cannot be expressed, but the reporter gene in yeast cells with specific CCAAT cis-acting elements can be expressed, indicating that the transcription factor can bind to CCAAT cis-...

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Abstract

The invention discloses a plant stress tolerance-related protein GmNF-YC9 as well as an encoding gene and an application thereof. As shown in experiments, in a drought tolerance test, the survival rate of T3 generation of homozygous transgenic plants obtained by transforming a recombinant expression vector pBI121-GmNF-YC9 of a DAN molecule expressed by 34th to 897th nucleotide sequences in a sequence 2 in a sequence table into arabidopsis is 90.2%, and the survival rates of wild plants and transgenic empty vector plants are respectively 27.2% and 28.4%; and in a salt-tolerant germination rate test, the germination rate of the T3 generation of homozygous transgenic plants is 90.4%, and the germination rates of the wild plants and the transgenic empty vector plants are respectively 65.8% and 66.2%. The GmNF-YC9 protein and the encoding gene thereof provided by the invention have great significance for improving the stress resistance of plants.

Description

technical field [0001] The present invention relates to a protein related to stress tolerance and its coding gene and application in the field of biotechnology, in particular to a plant stress tolerance related protein GmNF-YC9 and its coding gene and application. The protein GmNF-YC9 is derived from Soybean, has the ability to increase plant tolerance to drought and salt. Background technique [0002] Adversity stresses such as drought, high salinity and low temperature severely restrict the growth and development of soybean. Therefore, understanding the response and signal transduction mechanism of soybean to stress conditions and improving the stress resistance of soybean varieties have become one of the important tasks of soybean genetic research and variety improvement. [0003] Under adversity stress, plants will produce a series of responses, accompanied by many physiological, biochemical and developmental changes. Clarifying the response mechanism of plants to stre...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/63C12N5/10C12N1/21C12N1/19C12N7/01C12N15/82A01H5/00
CPCC07K14/415C12N15/8273
Inventor 马有志徐兆师郑炜君李连城陈明
Owner INST OF CROP SCI CHINESE ACAD OF AGRI SCI
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