Application of atsrt2 Gene in Improving Salt Tolerance of Plants

A plant and active technology, applied in the field of molecular genetics, can solve the problem of no AtSRT2 mediating plant salt tolerance, and achieve the effects of improving plant survival rate, salt tolerance, and seed germination rate.

Pending Publication Date: 2016-11-02
GUIZHOU INST OF PRATACULTURE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

AtSRT2 is a histone deacetylase, so far, there is no report on AtSRT2 mediating plant salt tolerance

Method used

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  • Application of atsrt2 Gene in Improving Salt Tolerance of Plants
  • Application of atsrt2 Gene in Improving Salt Tolerance of Plants
  • Application of atsrt2 Gene in Improving Salt Tolerance of Plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0038] Example 1 , seed germination period AtSRT2 expression under salt stress induce

[0039] 1. Extraction of Arabidopsis total RNA

[0040] The seeds of wild type Arabidopsis Columbia0 were treated with 70% (volume fraction) alcohol in an ultra-clean workbench for 3 minutes, and then washed twice with sterile water, each time for 1 minute, with 5‰ (mass volume ratio) sodium hypochlorite. After sterilizing for 10 minutes, wash twice with sterile water for one minute each time. The control group sowed sterile seeds in MS0 liquid medium with a pipette gun, and transferred them to the tissue culture room after vernalization at 4°C for 3 days. Samples were taken at 0h, 6h, 12h, 24h, 36h, and 48h. At the same time, the salt-treated group used a pipette gun to sow the sterile seeds in MS0 liquid medium containing 60mM NaCl, and transferred them to the tissue culture room after vernalization at 4°C for 3 days. , 60h, 72h were sampled.

[0041] The total RNA was ext...

Embodiment 2

[0050] Example 2 , Protoplast subcellular localization

[0051] 1. Construction of subcellular localization vector

[0052] Specific primers AtSRT2-F2 and AtSRT2-R2 were designed according to the CDS sequence of the AtSRT2 gene provided on the TAIR website to amplify the full length of the AtSRT2 gene (remove the stop codon). Use Infusion enzyme to connect the target fragment to the BamHI-digested subcellular localization vector 16318GFP. The ligation reaction system is: 2 μl Infusion HD Enzyme Premix, 5 μl 16318GFP digestion product, 3 μl PCR amplification product, and react at 50°C for 15 minutes . After the ligation reaction, heat shock transformed E. coli Top10. PCR detection of recombinant plasmids. Positive clones were sequenced. The plasmid was extracted to obtain the recombinant plasmid 116318-AtSRT2-GFP.

[0053] The primer sequences are:

[0054] 16318-AtSRT2-GFP-F:5'-TATCTCTAGA GGATCC ATGCTTTCTATGAACATGAGAAG-3' (the underlined part is the recognition si...

Embodiment 3

[0075] Example 3 、 AtSRT2 Gene cloning and obtaining overexpression lines

[0076] 1. Cloning of AtSRT2 gene and construction of recombinant expression vector pBI121-AtSRT2

[0077] The wild-type Arabidopsis Columbia0 at the adult plant stage was used as the experimental material, and the total RNA was extracted by the TRIzol method. The qualified RNA was reverse-transcribed into cDNA with the reverse transcription kit of TaKaRa Company, and stored at -20°C for future use. According to the TAIR website provided AtSRT2 Design specific primers for the CDS sequence of the gene AtSRT2 -F2 and AtSRT2 -R2, amplified AtSRT2 Gene full length. The reaction system includes 2×GC Buffer 25μl, dNTP Mix 4μl, upstream and downstream primers 1μl, Primerstar 0.3μl, ddH 2O 16.7 μl, template cDNA 2 μl (the above PCR reaction reagents are from TaKaRa company PCR amplification kit). The PCR amplification program was pre-denaturation at 94°C for 10 min, denaturation at 94°C for 30 ...

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Abstract

The invention discloses an AtSRT2 protein, which is characterized in that it is a protein having the amino acid sequence shown in SEQ ID NO: 1 in the sequence table, or the amino acid residue sequence of SEQ ID NO: 1 is passed through more than one amino acid residue sequence. A protein derived from SEQ ID NO: 2 which has the same activity as the amino acid residue sequence of SEQ ID NO: 1 without substituting, deleting or adding groups. Introduce the coding sequence of the AtSRT2 protein provided by the present invention into the target plant by genetic engineering to obtain a transgenic plant with higher salt tolerance than the target plant, so that the salt tolerance of the plant is improved, which is mainly reflected in: Under salt treatment conditions, compared with the wild type, the seed germination rate of the plants transferred with the AtSRT2 protein coding sequence was significantly improved; the salt tolerance was enhanced, the plant growth was better, and the root and leaf area were significantly higher than the wild type. Compared with the wild type; the survival rate of the plants was significantly improved. The present invention plays an important role in the breeding of plants for salt tolerance and enhanced salt tolerance.

Description

technical field [0001] The invention relates to the field of molecular genetics, in particular to an AtSRT2 protein and its coding sequence and application. Background technique [0002] Soil salinization has become a major environmental and resource problem worldwide that threatens human existence. According to incomplete statistics from the Food and Agriculture Organization of the United Nations, the area of ​​salinized land in the world is as high as 954 million hm 2 , seriously restricting the sustainable development of agriculture and animal husbandry. The area of ​​various types of salinized land in my country is equivalent to 1 / 4 of the existing cultivated land. In recent years, with the continuous expansion of the land salinized area, the national soil salinized area has reached 12 million hm 2 , is a big country with saline land in the world (Li Bin et al., 2005). Salt stress severely inhibits plant growth and development, and is an important environmental stress...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/82C12N15/29A01H5/00
Inventor 钟理陈明杨春燕马有志吴佳海
Owner GUIZHOU INST OF PRATACULTURE
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