Plant heat resistance related protein TaXPD and coding gene and application thereof

A plant and coding technology, which is applied in the field of plant heat resistance-related protein TaXPD and its coding gene and application field, can solve the problems of harm, yield decline, quality reduction, etc., and achieve the effect of increasing heat resistance

Active Publication Date: 2017-01-11
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The suitable temperature for wheat grain filling period is 20-24°C, and the relative humi

Method used

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  • Plant heat resistance related protein TaXPD and coding gene and application thereof
  • Plant heat resistance related protein TaXPD and coding gene and application thereof
  • Plant heat resistance related protein TaXPD and coding gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0077] Cloning of the coding gene of embodiment 1, protein TaXPD

[0078] The gene encoding the protein TaXPD, ie the TaXPD gene, was cloned from wheat. Specific steps are as follows:

[0079] 1. Use TRNzol Total RNA Extraction Kit to extract total RNA from fresh wheat leaves, and then use M-MLV reverse transcriptase to reverse transcribe the first-strand cDNA.

[0080] 2. Artificially synthesized primers F: 5'-ATGAAGTTCGATCTTGAAGG-3' and R: 5'-TCACATCTCCATGGCGTCCC-3'.

[0081] 3. After completing steps 1 and 2, use the cDNA extracted in step 1 as a template and use F and R as primers for PCR amplification to obtain a double-stranded DNA molecule of about 2300 bp.

[0082] 4. Ligate the double-stranded DNA molecule with pEASY-Blunt Cloning Vector to obtain recombinant plasmid A.

[0083] According to the sequencing results, the recombinant plasmid A contains the DNA molecule shown in sequence 1 in the sequence listing (hereinafter referred to as TaXPD gene), and expresses t...

Embodiment 2

[0084] Embodiment 2. Real-time quantitative detection of the relative expression of TaXPD gene in wheat samples under the condition of high temperature treatment for different time

[0085] Three replicates were performed, and the steps for each replicate were as follows:

[0086] (1) Get samples

[0087] The 10-day-old wheat seedlings were treated at 22° C. for 3 hours, and then the leaves were removed and stored in liquid nitrogen to obtain sample 1 (as a control).

[0088] Wheat seedlings grown to 10 days were treated at 40°C for 1 hour, and then the leaves were taken out and stored in liquid nitrogen to obtain sample 2.

[0089] According to the above method, replace 1h with 2h, 3h, 6h, and 12h respectively, and keep other steps unchanged, and obtain sample 3, sample 4, sample 5, and sample 6 in sequence.

[0090] (2) Real-time quantitative detection of the relative expression of TaXPD gene in wheat samples processed for different times under high temperature conditions ...

Embodiment 3

[0096] Embodiment 3, the acquisition and identification of transgenic Arabidopsis

[0097] 1. Acquisition of recombinant plasmids pB2GW7-TaXPD and GV3101 / pB2GW7-TaXPD

[0098]1. Acquisition of recombinant plasmid pB2GW7-TaXPD

[0099] Using Gateway technology to construct an overexpression vector, the specific steps are as follows:

[0100] (1) Acquisition of the target gene

[0101] Primers were designed according to the open reading frame sequence of the TaXPD gene as follows:

[0102] TaXPD-TOPOF: 5'-CACCATGAAGTTCGATCTTGAAGG-3'

[0103] TaXPD-TOPOR: 5'-TCACATCTCCATGGCGTCCC-3'.

[0104] TRNzol Total RNA Extraction Kit was used to extract total RNA from fresh wheat leaves, and then M-MLV reverse transcriptase was used to reverse transcribe the first-strand cDNA to obtain a cDNA solution. The DNA concentration in the cDNA solution was 1000 ng / μL.

[0105] The cDNA solution is used as a template, and TaXPD-TOPOF and TaXPD-TOPOR are used as primers for PCR amplification to ...

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Abstract

The invention discloses plant heat resistance related protein TaXPD and coding gene and application thereof. The protein provided by the invention is a1) or a) or a3: a) amino acid sequence and is a protein as shown in a sequence 2 in a sequence table; a) is fusion protein which is obtained by connecting label to an end N or/and an end C of the protein as shown in the sequence 2 in the sequence table; and a3) is protein which is related to plant heat resistance and is obtained by substitution and/or deficiency and/or addition of one or several amino acid residues of the protein as shown in a1) or a2). Experiments prove that the coding gene of the protein is guided into an arabidopsis mutant uvh6-1 to obtain a transgenic plant of which the survival rate is increased and/or the extension length of a hypocotyledonary axis is increased and/or the ion leakage rate is reduced. Therefore, the protein provided by the invention has important theoretical significance and practical value on cultivation of plants of which heat resistance are improved.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to plant heat-resistant related protein TaXPD and its coding gene and application. Background technique [0002] With the intensification of the greenhouse effect, the threat of high temperature stress to crops is becoming more and more serious. It is estimated that during the dry season, every 1°C increase in nighttime temperature will reduce rice yield by 10%. The increase in temperature has also had a serious impact on major crops such as wheat, corn, and barley. During the 20 years from 1981 to 2002, the increase in temperature has reduced the production of wheat, corn, and barley by about 40 million tons per year worldwide. Equivalent to an economic loss of $5 billion. In the summer of 2013, extreme high temperature weather occurred in the eastern and southern parts of my country, which seriously affected various crops including wheat, corn and rice. [0003] Wheat is the main f...

Claims

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Application Information

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IPC IPC(8): C07K14/415C07K19/00C12N15/29C12N15/62C12N15/82A01H5/00
Inventor 孙其信胡兆荣马俊华郑美倪中福姚颖垠彭惠茹辛明明李慧敏
Owner CHINA AGRI UNIV
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