CN112280783B

CN112280783B - CvHSF30-2 gene and application of coded protein in improving high temperature tolerance of plants or cells

Info

Publication number: CN112280783B
Application number: CN202011158291.9A
Authority: CN
Inventors: 明凤; 王锐; 彭思远; 毛婵娟; 蒋昌华; 莫健彬; 奉树成
Original assignee: SHANGHAI BOTANICAL GARDEN; Shanghai Normal University
Current assignee: SHANGHAI BOTANICAL GARDEN; Shanghai Normal University
Priority date: 2020-10-26
Filing date: 2020-10-26
Publication date: 2023-11-14
Anticipated expiration: 2040-10-26
Also published as: CN112280783A

Abstract

The invention discloses a nucleotide sequence which is selected from the following sequences: a nucleotide sequence shown as SEQ ID NO.1 or SEQ ID NO. 5; nucleotide sequence derived from the nucleotide sequence shown in SEQ ID No.1 or SEQ ID No.5 by substitution, deletion or addition of one or several nucleotides; a nucleotide sequence having at least 80% homology with SEQ ID No.1 or SEQ ID No. 5. The nucleotide sequence can improve the high temperature tolerance of plants or cells.

Description

CvHSF30-2 gene and application of coded protein in improving high temperature tolerance of plants or cells

Technical Field

The invention belongs to the technical fields of molecular biology and genetic engineering, and particularly relates to a gene for improving high-temperature tolerance of plants and application thereof.

Background

As the phenomenon of the greenhouse effect is increasingly obvious, the global air temperature is continuously increased, and the high temperature in summer becomes a main environmental factor for restricting the growth and development of plants, the growth and development of the plants face a serious challenge of high-temperature adversity. The heat shock transcription factor (heat shock transcription factor, HSF) is an important transcription factor of plants, and is combined with cis-acting elements of a promoter of a heat shock response related gene to accurately regulate the space-time expression of a downstream gene and regulate the heat shock response of the plants. Heat shock transcription factor (heat shock factor, HSF) is a transcription factor found earlier in eukaryotes and is ubiquitous in bacteria, algae, animals, plants. There are more genes encoding HSF in the plant genome than other eukaryotes such as yeast, drosophila and animals (1 in yeast, nematodes, 4 in vertebrates), and the number of HSF family members in many plant species exceeds 20 (21 in arabidopsis), which may be due to gene replication and functional differentiation during plant evolution.

HSF heat induction is a characteristic of plants that is not found in yeast and animals. Members of Class A1 in Arabidopsis (HSFA 1a/HSFA1b/HSFA1 d/HSFA1 e) play a major role in activating transcription of heat-inducible genes (including HsfA 2), which trigger a transcription cascade consisting of a number of transcription factors when plants are subjected to heat shock. In tomato, hsfA1 is the main regulator of plant heat resistance. HsfA1 controls the protective pathway of heat stress. In wheat, taHsfA6f and TaHsfC2a also protect plants from heat stress.

With the improvement of living standard, ornamental plants in gardens are attracting more and more attention, and the research of genetic mechanisms of ornamental plants on temperature stress resistance has wide application prospect. Accordingly, those skilled in the art have been working to find genes that increase the high temperature tolerance of plants and their uses.

Disclosure of Invention

In view of the shortcomings in the prior art, the invention provides a gene for improving high temperature tolerance of plants or cells and application thereof.

In a first aspect the invention provides a gene for increasing the high temperature tolerance of a plant or cell, in a specific embodiment the nucleotide sequence is selected from the group consisting of:

1) A nucleotide sequence shown as SEQ ID NO.1 or SEQ ID NO. 5;

2) Nucleotide sequence derived from the nucleotide sequence shown in SEQ ID No.1 or SEQ ID No.5 by substitution, deletion or addition of one or several nucleotides;

3) A nucleotide sequence having at least 80% homology with SEQ ID No.1 or SEQ ID No. 5.

In a second aspect the invention provides an amino acid sequence which increases high temperature tolerance in a plant or cell, in one embodiment the amino acid sequence is selected from the group consisting of:

1) An amino acid sequence shown in SEQ ID NO. 2;

2) An amino acid sequence derived from the amino acid sequence shown in SEQ ID NO.2 by substitution, deletion, or addition of one or more amino acids;

3) An amino acid sequence having at least 80% homology with SEQ ID NO. 2.

In a third aspect the invention provides the use of a nucleotide sequence as described above or an amino acid sequence as described above for increasing the high temperature tolerance of a plant or cell.

Further, the cell is a prokaryotic cell or a eukaryotic cell.

Further, the prokaryotic cells include E.coli; eukaryotic cells include yeast.

In a fourth aspect the present invention provides a method of increasing the high temperature tolerance of a plant comprising introducing into the plant of interest a nucleotide sequence as described above.

Further, the nucleotide sequence as described above is ligated to a plant expression vector, and then the recombinant plant expression vector is transferred into a target plant, and a plant with high temperature tolerance is obtained by screening.

In a fifth aspect the invention provides a method of improving the high temperature resistance of a cell comprising introducing into a cell of interest a nucleotide sequence as described above.

Further, the nucleotide sequence as described above is ligated to an expression vector, and then the recombinant expression vector is transferred into a target cell, and a cell with high temperature resistance is obtained by screening.

Further, the cells include E.coli and yeast.

The heat shock transcription factor CvHSF30-2 cloned from Clematis provided by the invention is transferred into escherichia coli and yeast to improve the high temperature stress tolerance of a transgenic strain, so that the heat shock transcription factor CvHSF30-2 can improve the temperature resistance of prokaryotes and lower eukaryotes. Meanwhile, the tolerance of the Nicotiana benthamiana to temperature is influenced after the VIGS in the Nicotiana benthamiana TRV-NBHSF30-2, and the fact that the heat shock transcription factor and the homologous sequence thereof can indeed influence the high-temperature tolerance of plants is also shown.

Drawings

FIG. 1 shows the growth curve at 37℃of E.coli transformed with CvHSF30-2 and an empty control in example 2 of the present invention.

FIG. 2 is a graph showing the growth of E.coli transformed with CvHSF30-2 and an empty control at 50℃in example 2 of the present invention.

FIG. 3 shows colony growth at 37℃and 50℃for the empty control of E.coli transfected with CvHSF30-2 in example 2 according to the present invention.

FIG. 4 shows the 28℃growth curve of yeast AH109 transformed with CvHSF30-2 and a blank in example 3 of the present invention.

FIG. 5 shows the growth curves of yeast AH109 transformed with CvHSF30-2 and the blank at 37℃and 42℃in example 3 of the present invention. Wherein FIG. 5a is a growth curve of 37℃culture and FIG. 5b is a growth curve of 42℃culture.

FIG. 6 shows growth of heat-treated colonies of yeast AH109 transformed with CvHSF30-2 and blank at 37℃and 42℃in example 3 of the present invention.

FIG. 7 shows qPCR results of the tranformed TRV: nbHSF30-2 tobacco in example 4 of the present invention.

FIG. 8 is a heat treatment phenotype of the tran-TRV: nbHSF30-2 tobacco in example 4 of the present invention.

FIG. 9 shows the NBT and DAB staining results of tobacco leaves with a heat treatment phenotype of NbHSF30-2 tobacco in example 4 of the present invention.

Detailed Description

The invention will be further illustrated with reference to the following examples, which are to be understood as illustrative only and are not intended to limit the scope of the invention.

The experimental procedure, which does not specify specific conditions in the following examples, is generally followed by conventional conditions, such as molecular cloning by Sambrook et al: conditions described in the laboratory Manual (New York: cold Spring Harbor Laboratory Press, 1989) or as recommended by the manufacturer. The reagents used, unless otherwise specified, are commercially available or publicly available.

In the present invention, various vectors known in the art, such as commercially available vectors including plasmids and the like, may be used.

The clematis has a heat-resistant variety and a thermolabile variety, and researches show that the CfHSF30-2 gene in the clematis is expressed by heat shock induction, but under the condition of high temperature stress, the CfHSF30-2 gene is only strongly expressed in the thermoresistant clematis variety, but is weakly expressed or even not expressed in the thermolabile variety, and whether the gene is expressed or not is consistent with the field heat resistance difference observation result of the clematis variety.

EXAMPLE 1 cloning of the clematis CvHSF30-2 Gene

Extracting total RNA of tender leaves of the clematis vitis vinifera which is subjected to heat shock treatment, wherein an extraction kit is RNAplant (commercially available), and carrying out reverse transcription on the total RNA into cDNA by using a reverse transcription kit (commercially available). The primer is designed according to the sequencing result of the transcriptome, the primer sequences are shown as SEQ ID NO.3 and SEQ ID NO.4, and a 927bp band is amplified from the clematis vitis vinifera cDNA by adopting an RT-PCR method. And (3) recovering a PCR product to obtain the CvHSF30-2 gene, wherein the nucleotide sequence of the gene is shown as SEQ ID NO.1, the amino acid sequence coded by the nucleotide sequence is shown as SEQ ID NO.2, the gene consists of 308 amino acid residues, and the molecular weight of the gene is 35.21 kilodaltons.

EXAMPLE 2 Leptoradix Clematidis Botrytis CvHSF30-2 prokaryotic expression for improving heat resistance of escherichia coli

(1) The open reading frame of the CvHSF30-2 gene was operably linked to a prokaryotic expression vector (PET-28 a) to form a prokaryotic expression vector containing the gene PET-28a-CvHSF30-2.

(2) The PET-28a-CvHSF30-2 expression vector is transferred into a prokaryotic escherichia coli DH5 alpha, and the plasmid PET-28a-CvHSF30-2 is extracted from the obtained positive clone.

(3) Plasmid PET-28a-CvHSF30-2 was transformed into E.coli BL (DE 3) for expression, namely BL (DE 3) containing plasmid PET-28a-CvHSF30-2 was cultured overnight (37 ℃ C., 220 rpm) in liquid LB (Kan+ final concentration of 100 mg/L) with shaking, and the OD of the bacterial solution was increased ₆₀₀ Value is adjusted to OD ₆₀₀ =1, bacterial liquid was prepared according to 1:100 is inoculated into LB (Kan+) liquid culture medium, shaking is carried out at 37 ℃, OD value is measured by taking bacterial liquid every 1h, culturing is carried out for 10h, and a growth curve is drawn. BL21 (DE 3) transferred into the empty PET-28a vector was used as a control to examine whether the introduction of CvHSF30-2 would affect the normal growth of E.coli. As shown in FIG. 1, the introduction of CvHSF30-2 did not affect the normal growth of prokaryotes BL (DE 3), and the growth trend was consistent with that of the transfer-in no-load.

(4) Taking the above-mentioned regulation OD ₆₀₀ 1 bacterial liquid, according to 1:100 (LB, kan+), 20ml, 200ul, placing the bacterial liquid at 50deg.C, shaking at high temperature, taking bacterial measurement OD value per hour,culturing for 3h. BL21 (DE 3) transferred into the empty PET-28a vector is used as a control to check the growth condition of the strain transferred into the CvHSF30-2 under the high temperature stress of 50 ℃.

(5) Taking the above-mentioned regulation OD ₆₀₀ As 1-fold bacterial solution, Z-streaked on LB plates (Kan+) and cultured overnight at 37℃and 50℃respectively. BL21 (DE 3) transferred into the empty PET-28a vector is used as a control, and the growth condition of the escherichia coli introduced with the CvHSF30-2 under the high temperature stress of 50 ℃ is observed.

As shown in FIGS. 2 and 3, both the growth curve and colony growth of the transformed CvHSF30-2 strain were shown to significantly improve the heat resistance of the prokaryote BL (DE 3) by CvHSF30-2, whether cultured in liquid medium or solid medium.

EXAMPLE 3 enhancement of Yeast Heat resistance by eukaryotic expression of Leptoradix Clematidis CvHSF30-2

The open reading frame of the CvHSF30-2 gene is operably linked to a eukaryotic expression vector to form a eukaryotic expression vector containing the gene, and the expression vector is transferred into eukaryotic yeast AH109, as shown in FIG. 4, wherein CvHSF30-2 does not affect the normal growth of yeast AH 109: as shown in FIG. 5a, FIG. 5b and FIG. 6, compared with the control, the strain transformed with CvHSF30-2 showed that the strain transformed with CvHSF30-2 can obviously improve the heat resistance of eukaryotic yeast AH109 under the conditions of bacterial growth and colony growth at 37 ℃ and 42 ℃.

(1) The open reading frame of the CvHSF30-2 gene is operably linked to a eukaryotic expression vector (BD) to form a prokaryotic expression vector BD-CvHSF30-2 containing the gene.

(2) The vector in step (1) was transferred into yeast (AH 109), and the yeast containing the expression vector was shake-cultured in liquid SD/-Trp (final concentration 100 mg/L) (28 ℃,220 rpm).

(3) Taking the yeast liquid cultured in the step (2), and regulating to OD ₆₀₀ The value was 0.05, and the culture was performed by shaking (28 ℃,220 rpm), and the OD of the bacterial liquid was measured every two hours ₆₀₀ Values, for a total of 32h, were plotted for yeast growth. AH109 transferred into empty BD vector was used as a control to see if introduction of CvHSF30-2 would affect normal growth of yeast. As shown in FIG. 4, the introduction of CvHSF30-2 did not affect the normal growth of eukaryotic yeast (AH 109), its growth trend and transfer-in-airThe growth trend of the load is consistent.

(4) Taking the yeast liquid cultured in the step (2), and regulating to OD ₆₀₀ The value is 0.05, the culture is carried out for 0, 6, 12, 18 and 24 hours by shaking table at 37 ℃ and 42 ℃ and 220rpm, and OD is measured ₆₀₀ Values, a growth curve is plotted. The growth of strain CvHSF30-2 was examined under high temperature stress at 42℃with AH109 transferred into empty BD vector as a control.

(5) Taking the yeast liquid cultured in the step (2), and regulating to OD ₆₀₀ Plates were spotted at a value of 1, diluted by various factors and grown overnight at 37℃and 42 ℃. The growth of strain CvHSF30-2 was examined under high temperature stress at 42℃with AH109 transferred into empty BD vector as a control.

As shown in FIGS. 5a, 5b and 6, both the growth curve and colony growth of the yeast strain transformed with CvHSF30-2 were shown to significantly improve the heat resistance of eukaryotic yeast AH109, whether cultured in liquid medium or in solid medium.

EXAMPLE 4 VIGS down-regulating tobacco NbHSF30-2 homologous Gene to decrease tobacco Heat resistance

(1) Selecting a sequence with highest homology with a clematis CvHSF30-2 gene in Nicotiana benthamiana, nbHSF30-2 (shown as SEQ ID NO. 5), taking the sequence as a template, designing a VIGS primer, and connecting a part of the sequence (shown as SEQ ID NO. 6) to a TRV2 virus vector to form a recombinant vector.

(2) The recombinant vector is transformed into agrobacterium tumefaciens GV3101, and meanwhile, the virus TRV1 vector is also transformed into the agrobacterium tumefaciens GV3101, two GV3101 bacterial liquids are obtained through culture, then the Nicotiana benthamiana is injected together, and the NbHSF30-2 gene of the tobacco is silenced in a virus-induced gene silencing (Virus induced gene silence, VIGS) mode.

(3) After 2 weeks of injection, qPCR was performed to verify the down-regulation of tobacco NbHSF 30-2.

(4) The qPCR-verified down-regulated tobacco of NbHSF30-2 was subjected to 42℃heat treatment (heat treatment in a 42℃light incubator for 1 h), and the phenotype was examined to further verify the heat-resistant function of CvHSF30-2.

(5) After heat treatment, TRV2-HSF30-2 showed significant loss of water and wilting, while the phenotype of the control TRV2-GFP was not significantly different from that before treatment. The activity of superoxide dismutase (SOD) was measured by performing a blue tetrazolium (NBT) staining method on TRV2-HSF30-2 and TRV2-GFP tobacco leaves, and the activity of Peroxidase (POD) in cells was measured by a diaminobenzidine staining method (DAB).

The steps (2), (3), (4) and (5) are all performed by using TRV2-GFP as a parallel control.

As shown in FIG. 7, the NbHSF30-2 gene in tobacco was smoothly reduced by VIGS. As shown in FIG. 8, PDS is a phytoene dehydrogenase gene, and in this experiment, TRV-PDS gene was silenced to show white color, indicating that the gene silencing effect was good. After 42℃heat treatment, tobacco plants in which the VIGS down-regulates the tobacco NbHSF30-2 homologous gene exhibited leaf sagging and atrophy compared with TRV2-GFP, indicating that down-regulating tobacco NbHSF30-2 reduced tobacco heat resistance. As shown in FIG. 9, after the VIGS down-regulates the tobacco NbHSF30-2, the SOD and KOD levels of the tobacco are increased, namely the active oxygen level is obviously increased, the damage to the tobacco caused by heat stress is larger, and the heat resistance of the tobacco is reduced.

Sequence listing

<110> Shanghai university; shanghai plant garden

<120> use of CvHSF30-2 gene and encoded protein to increase high temperature tolerance in plants or cells

<130> CN015-20007PICN

<160> 6

<170> SIPOSequenceListing 1.0

<210> 1

<211> 927

<212> DNA

<213> Clematis (Clematis florida thunder.)

<400> 1

atgatcaagg aggaagatgt tgtggaagtt atggatcttg atggcgcttc ttcgtcaatg 60

ccactgtatc cccagccagt ggaagggatg catgaggtag gccctccacc atttctgacg 120

aagatattcg aaatggtgga agacccggca actgattcaa ctgtttcatg gagcagagca 180

caaaacagtt ttattgtttg ggacttgccc aagctttcaa ccactcttct tccaagatac 240

ttcaagcaca gtaacttctc tagctttgta aggcaactca acacatatgg gtttaggaag 300

gttgatccgg acagatggga gttcgctaat gaagggtttc tcggagggca gaagcatttg 360

ttgaaaaaca ttaagaggag aagaaatgtc tcagcctctc acaccatgca acagggattt 420

gaaccttgcg ttgacttggg ggaatatgga ctggaagctg agatgagcac tctgaagagg 480

gatcgcaact atttgatgtc tgagattatg aagctaagag agcatcagat agactcccgt 540

gaaaaagtta tagcgattgg tgaacgcttg cgagcaaccg aggggaagaa ccagcgtatg 600

gggaatttcc tggcgagagc tatcaagaat ccagaatttc tccagaaact ggaggaggtg 660

aaggaactac cggattttga aattggaagg aaaaggaaac tggcagacaa ttatgtaatg 720

gggaacttgc aagctgtgga agaagtgatc gattctgaca tagagacgct attctctcca 780

aacggaactg aaactgaaat ccttcagctg gagactggtt cagctaactc tatgttgtgg 840

gaagatttgg taaatgacga gttgattgcg tgtgcggaag tggatgatct gttcaaggag 900

atgggttgcc ttgggccaaa gccgtga 927

<210> 2

<211> 308

<212> PRT

<213> Clematis (Clematis florida thunder.)

<400> 2

Met Ile Lys Glu Glu Asp Val Val Glu Val Met Asp Leu Asp Gly Ala

1 5 10 15

Ser Ser Ser Met Pro Leu Tyr Pro Gln Pro Val Glu Gly Met His Glu

20 25 30

Val Gly Pro Pro Pro Phe Leu Thr Lys Ile Phe Glu Met Val Glu Asp

35 40 45

Pro Ala Thr Asp Ser Thr Val Ser Trp Ser Arg Ala Gln Asn Ser Phe

50 55 60

Ile Val Trp Asp Leu Pro Lys Leu Ser Thr Thr Leu Leu Pro Arg Tyr

65 70 75 80

Phe Lys His Ser Asn Phe Ser Ser Phe Val Arg Gln Leu Asn Thr Tyr

85 90 95

Gly Phe Arg Lys Val Asp Pro Asp Arg Trp Glu Phe Ala Asn Glu Gly

100 105 110

Phe Leu Gly Gly Gln Lys His Leu Leu Lys Asn Ile Lys Arg Arg Arg

115 120 125

Asn Val Ser Ala Ser His Thr Met Gln Gln Gly Phe Glu Pro Cys Val

130 135 140

Asp Leu Gly Glu Tyr Gly Leu Glu Ala Glu Met Ser Thr Leu Lys Arg

145 150 155 160

Asp Arg Asn Tyr Leu Met Ser Glu Ile Met Lys Leu Arg Glu His Gln

165 170 175

Ile Asp Ser Arg Glu Lys Val Ile Ala Ile Gly Glu Arg Leu Arg Ala

180 185 190

Thr Glu Gly Lys Asn Gln Arg Met Gly Asn Phe Leu Ala Arg Ala Ile

195 200 205

Lys Asn Pro Glu Phe Leu Gln Lys Leu Glu Glu Val Lys Glu Leu Pro

210 215 220

Asp Phe Glu Ile Gly Arg Lys Arg Lys Leu Ala Asp Asn Tyr Val Met

225 230 235 240

Gly Asn Leu Gln Ala Val Glu Glu Val Ile Asp Ser Asp Ile Glu Thr

245 250 255

Leu Phe Ser Pro Asn Gly Thr Glu Thr Glu Ile Leu Gln Leu Glu Thr

260 265 270

Gly Ser Ala Asn Ser Met Leu Trp Glu Asp Leu Val Asn Asp Glu Leu

275 280 285

Ile Ala Cys Ala Glu Val Asp Asp Leu Phe Lys Glu Met Gly Cys Leu

290 295 300

Gly Pro Lys Pro

305

<210> 3

<211> 23

<212> DNA

<213> Artificial sequence (Artificial Sequence)

<400> 3

atgatcaagg aggaagatgt tgt 23

<210> 4

<211> 24

<212> DNA

<213> Artificial sequence (Artificial Sequence)

<400> 4

cggctttggc ccaaggcaac ccat 24

<210> 5

<211> 2343

<212> DNA

<213> Benshi tobacco (Nicotiana benthamiana)

<400> 5

aacgaaaatg cctcgttggc gcaaatcaag cgttccatga ctttctcgcg tcctctagtt 60

tccaatttcc accgtttggt gccagtcaat ctctccccta ctactagtcc ctcctccttt 120

ctctttagct cactagcttg ttagtttatc gagaaaacgg tgcgattgaa tggacggagt 180

tcatgagaac acggtggtag ggaattcgcc gcctcctttt ctgagcaaga cgtatgacat 240

ggttgatgat tcgtcgacag attctgtggt gtcgtggagt aagagtcaca acagctttgt 300

tgtgtggaac gtgccggagt ttgctagaga tgttttgccc aagtatttca agcacaataa 360

tttctccagc ttcgtcagac agttgaatac ttacggtttc aggaaggttg atccggatcg 420

ctgggaattt gcaaacgagg gatttcttag aggccacaaa caccagttga agagtataag 480

caggcgaaaa ccctctcagg tgcagggtca tcagcaaaca acctcccaag tgcaaaatgc 540

atctgttggg tcttgtgttg aagttgggaa gttcggaaca gaggatgagg tggaaaggct 600

taaaagagat aaggatgttc ttatgcagga gctggttgag ttgaaacagc agcagcaggc 660

aacggatcac caaatgcaga ctgttgggca gcgtgtccaa ttaatggaga aaaggcaaca 720

gcaaatgatg tcattccttg ccaaggccat gcagagccct ggctttgtag cccaactggt 780

acatcaacag aatgggaata acaggcgtat tgcgggaatg aataagaaaa ggagacttcc 840

tgaccaggat gaagaaaatg ttgcaggcaa tcctgttaat atgtcgcgag atggacggat 900

agtcaggtat caaccttcaa tgaatgaggc agcaaaagca ttgctgcggc agattctgaa 960

aataaataca cctgggaggc tggacaatag acttaagaat acaaatggtc tgttgactga 1020

taatcccctc tcccccaaga atgcacttag tggtgataca tccagccgta tttctggggt 1080

taccctctct gaagtattgt caatgtcttc acagtctcat ataatgtcag attcaggatt 1140

tccattcagc tcttgttcat ctgttatgtc tgagagccca tcttcaacta ctgtggttcc 1200

tggggaggcc aaagttccta cgttatgtga aacggatgct ctcaactctc tcacagacca 1260

tgatttgcct gaattttctc aagaacaaag gattaataga caggaaactt ttggaataac 1320

tggccttaag aggtctgaga ctggaaatgt gccatatatt gacacgcaag cctttttgga 1380

tggtgtaaca gcaattgtcc ctgatggatt ttcggcagac acccatgtcg atgtcctgct 1440

agatgatgtg cctaagcttc cggcgattaa tgatattttc tgggaccaga ttctttctgc 1500

aagccctctt actggggata tggatgaaat tggtattgaa gatggcttag aaaaggaaga 1560

ggattttcta ggagttcagg agaatgattt ggataaacta aaccatatgt gtcatcttac 1620

tgaacagatg gggcttcttt catcagcggc ccagatttga tgatttcatg gattttattt 1680

gaacactgca acaacggagg tggacactct gaactggtgg gcattaacaa gcttgctatg 1740

aaggactgat cttcactact actaatgatg ttctgagcag tgaaaagggg accccaaaag 1800

ataaagggtt gtgtctgatc ttgtattagt cttttgatgc atatgtatgc agactttgta 1860

tggatattat ccagcacaac tgttaggtaa aaaaggaata ctaaaatggt gtttacttgt 1920

taagaaagta agaaagggca cggtagaaag ttgttgcaga ggatctgttt gtaaactttg 1980

ctgtttacct aagcaacgac tgaggatttt aatgaagatg caagctgttg tgtggatggc 2040

ttgctttttg tacagatata ttgaatggta tttgtttgtt atcgtgaggt ttgtattcta 2100

tgcagaggtt gcttcgagca gcttctccca ctaaatagat gaatcctctt tcccagcata 2160

ttatgaaaat tttccctttc tgcgtcgctt attgttgggc tgatgctgat aacaaggggt 2220

gttcataaaa attgaaaaat cgagccgaac caaaaatcag actaaaccga cgaaaaaaat 2280

cgttactttt tggtttggtt tcgaatttta aaaattgatc gaatttggtt tggtttggaa 2340

ttt 2343

<210> 6

<211> 170

<212> DNA

<213> Artificial sequence (Artificial Sequence)

<400> 6

cctccttttc tgagcaagac gtatgacatg gttgatgatt cgtcgacaga ttctgtggtg 60

tcgtggagta agagtcacaa cagctttgtt gtgtggaacg tgccggagtt tgctagagat 120

gttttgccca agtatttcaa gcacaataat ttctccagct tcgtcagaca 170

Claims

1. An isolated gene, wherein the nucleotide sequence of the gene is shown in SEQ ID NO. 1.

2. A protein is characterized in that the amino acid sequence of the protein is shown as SEQ ID NO. 2.

3. Use of an isolated gene according to claim 1 or a protein according to claim 2 for increasing the high temperature tolerance of a plant or a cell, wherein the plant is clematis and the cell is escherichia coli or yeast.

4. A method for improving high temperature tolerance of a plant, comprising introducing the isolated gene of claim 1 into a plant of interest; wherein the plant is clematis terniflora.

5. The method for improving high temperature tolerance of a plant according to claim 4, wherein the isolated gene according to claim 1 is ligated to a plant expression vector, and then the recombinant plant expression vector is transferred into a target plant, and a plant having high temperature tolerance is obtained by screening.

6. A method for improving high temperature resistance of a cell, comprising introducing the isolated gene of claim 1 into a target cell; wherein the cell is Escherichia coli or yeast.

7. The method for improving high temperature resistance of cells according to claim 6, wherein the isolated gene according to claim 1 is ligated to an expression vector, and then the recombinant expression vector is transferred into a target cell, and the cells having high temperature resistance are obtained by screening.