Abiotic stress response related protein, and encoding gene and application thereof

A biological, protein technology, applied in the field of protein

Active Publication Date: 2015-05-13
BEIJING DABEINONG BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] So far, many genes related to plant salt tolerance have been cloned and studied, including genes encoding various organic solute synthases: such as the P5cs gene involved in proline synthesis, genes related to betaine syn...

Method used

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  • Abiotic stress response related protein, and encoding gene and application thereof
  • Abiotic stress response related protein, and encoding gene and application thereof
  • Abiotic stress response related protein, and encoding gene and application thereof

Examples

Experimental program
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Effect test

no. 1 example

[0068] The first embodiment, construction of sorghum full-length cDNA library

[0069] Take salt-treated sorghum tissue to prepare total RNA, and after mRNA separation, dephosphorylation, decapping, and CAP Tag connection, the first strand of cDNA is synthesized from the purified mRNA, and then double-stranded DNA is synthesized. After digestion, the fragments were fractionated by agarose gel electrophoresis, and the fragments were harvested according to the following fragment sizes: 2-12kb, 1-2kb, 0.5-1kb, figure 1 Shown (Spe: spectinomycin gene; RB: right border; prCaMV35s: cauliflower virus promoter (SEQ ID NO: 3); Sb: sorghum library gene; Nos: the terminator of nopaline synthase gene (SEQ ID NO: 4); Bar: glufosinate resistance gene (SEQ ID NO: 5); LB: left border).

[0070] Then, the recombinant expression vector mixture was transformed into Escherichia coli XL1-Blue competent cells by electroshock method. The electric shock conditions were: 50 μL E. coli XL1-Blue compete...

no. 2 example

[0071] The second embodiment, Arabidopsis transformation of sorghum full-length cDNA library

[0072] 1. Extraction of full-length cDNA library plasmid

[0073] Scrape all the plaques of the above-mentioned sorghum library and culture them in 500 mL LB liquid medium containing spectinomycin (100 mg / L) at 37°C for 30 min. The plasmid was extracted by alkaline method in small tubes: the bacterial solution was centrifuged at 12000 rpm for 1 min, the supernatant was removed, and 100 μL of ice-cold solution I (25 mM Tris-HCl, 10 mM EDTA (ethylenediaminetetraacetic acid) was used to precipitate the bacterial cells. 50mM glucose, pH8.0) suspension; add 200μL of newly prepared solution II (0.2M NaOH, 1% SDS (sodium dodecyl sulfate)), invert the tube 5 times, mix, put on ice for 3-5min; add 150 μL of ice-cold solution III (3M potassium acetate, 5M acetic acid), mixed thoroughly immediately, placed on ice for 5-10min; centrifuged at 4°C and 12000rpm for 5min, took an appropriate amount...

no. 3 example

[0081] The third embodiment, Arabidopsis screening of sorghum full-length cDNA library

[0082] Will T 1 Seeds were suspended in 0.1% agarose solution and stored at 4°C for 2 days. Mix horse manure with vermiculite and subirrigate with water until moist, allowing the soil mixture to drain for 24 hours. The pretreated seeds were sown on the soil mixture and covered with a moisture hood for 4 days. Make the seeds germinate and keep the light intensity at 120-150μmol / m at constant temperature (22°C) and constant humidity (40-50%) 2 Cultivated under long-day light conditions. When the Arabidopsis grew to 7 days, the non-transgenic Arabidopsis was killed by foliar spraying with Baoshida (Bayer) solution at a dilution ratio of 1:400. When the remaining transgenic Arabidopsis thaliana grows to 12-14 days, water it to saturation first, and then stop watering; when it grows to 18-20 days, start to measure the water content, and when the water content drops to 25-45%. The transgeni...

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Abstract

The invention relates to an abiotic stress response related protein, and an encoding gene and application thereof. The abiotic stress response related protein comprises a protein (a) having an amino acid sequence shown as SEQ ID NO: 2 or a protein (b) which is formed by performing substitution and/or deletion and/or addition on one or more amino acids in the amino acid sequence of the protein (a), is derived from the protein (a), and has an abiotic stress resistance activity. According to the invention, the abiotic stress response related protein is firstly isolated from sorghum, the protein is particularly tolerant to salt stress, the salt-tolerated protein/gene HPS1 is tolerated to salt stress with the concentration of 450 mM, and the protein is very important to improvement, development and utilization of saline land resources, land pressure relief, reserved cultivated land increase and guarantee of food security.

Description

technical field [0001] The present invention relates to a protein related to abiotic stress response, its encoding gene and application, in particular to a salt-tolerant protein derived from sorghum, its encoding gene and application. Background technique [0002] According to incomplete statistics, at least 20% of the cultivated land and more than 50% of the irrigated land in the world are affected by salt stress to varying degrees, and salt stress, as a kind of abiotic stress, is the main limitation of plant growth and yield Sexual factors can cause a series of physiological and metabolic reactions in plants, resulting in plant death or yield reduction. Since most plants, especially crops, are sweet soil plants sensitive to salt stress, salt stress has seriously affected global food production. Practice has proved that improving saline soil is a complex, difficult, and time-consuming task. Therefore, revealing the mechanism of plants coping with salt stress and improving ...

Claims

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Application Information

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IPC IPC(8): C07K14/415C12N15/29C12N15/82A01H5/00
CPCC07K14/415C12N15/8205C12N15/8261
Inventor 姚琴芳王飞鹏李晓娇杨进孝吕玉平
Owner BEIJING DABEINONG BIOTECHNOLOGY CO LTD
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