CN109627303A - The gene of Radix Notoginseng pathogenesis-related proteins PnPR3 and its application - Google Patents
The gene of Radix Notoginseng pathogenesis-related proteins PnPR3 and its application Download PDFInfo
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- CN109627303A CN109627303A CN201811509493.6A CN201811509493A CN109627303A CN 109627303 A CN109627303 A CN 109627303A CN 201811509493 A CN201811509493 A CN 201811509493A CN 109627303 A CN109627303 A CN 109627303A
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- C12N15/8271—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance
- C12N15/8279—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance for biotic stress resistance, pathogen resistance, disease resistance
- C12N15/8282—Phenotypically and genetically modified plants via recombinant DNA technology with agronomic (input) traits, e.g. crop yield for stress resistance, e.g. heavy metal resistance for biotic stress resistance, pathogen resistance, disease resistance for fungal resistance
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Abstract
The invention discloses the gene of Radix Notoginseng pathogenesis-related proteins PnPR3 a kind of, nucleotide sequence encodes the protein of the amino acid sequence as shown in SEQ ID NO:2 as shown in SEQ ID NO:1；The present invention expresses that it in tobacco by being transferred to foreign gene PnPR3 into tobacco, and experiment shows the overexpression of PnPR3 gene, has certain inhibiting effect to typical root rot germ Fusarium solani；Gene of the invention can be transferred to the disease resistance that plant is improved in plant, and tobacco can be improved to the resistance of root rot pathogenic bacteria Fusarium solani, lay good working foundation for root rot Resistant breeding, have high economic benefit.
Description
Technical field
The present invention relates to the encoding gene of Radix Notoginseng pathogenesis-related proteins PnPR3 a kind of and its applications, including Radix Notoginseng course of disease phase
Close protein genePnPR3And its coding protein and carrier and plant cell containing the gene.
Background technique
Series of genes response and up-regulated expression, some of which base after Radix Notoginseng is infected by pathogenic bacteria, in plant
Cause is related to defense reaction, including pathogenesis-related proteins (PR) and antibacterial gene, anti-to improve by enhancing its protein active
Sick ability；PR albumen is considered to be induced by pathogen and abiotic stress；According to the structure and function of PR albumen, PR at present
Albumen is found in monocotyledon and dicotyledon and has been acknowledged by 17 families.In these PR albumen, PnPR3 egg
It is white to take part in carbohydrate metabolism process, the biological processes such as chitin catalytic process and cell wall bulky molecular catalysis process,
The albumen has the molecular functions such as chitinase activity and chitin binding activity.
Radix Notoginseng (Panax notoginseng) it is the distinctive traditional rare traditional Chinese medicine in China, for many years, scale kind
Plant is primarily present the problem of continuous cropping obstacle, the serious limitation by diseases such as root rot, powdery mildew, Northern leaf spots, wherein with root-rot
Disease is Major Diseases, and long-term disease incidence is 5%~20%, continues to plant serious reachable 80% or more on notoginseng soil, very
To total crop failure, butt rot will lead to after the onset of root rot, the withered and yellow wilting of aerial part has seriously affected the yield and medicinal material of Radix Notoginseng
Quality.The control means of root rot are mainly largely to apply based on antibiotic property pesticide at present, and the Harmless control of root rot is
Become the hot and difficult issue problem of technique for planting notoginseng research.Notoginseng root rot pathogen is more complicated, generally compound to invade
Dye, pathogenic bacteria include the Pseudomonas alba (pseudomonas sp.) in bacterium, the corrupted column spore bacterium in fungi
(Cylindrocarpon destructans, C. didynum), sickle-like bacteria (Fusarium solani, Fusarium
Solani f. sp. Radicicola, F. oxysporum Schlecht., F. scirpi lamb.) etc., usually with bad
In damage column spore bacterium, sickle-like bacteria or pseudomonad based on certain one kind, several pathogen Combined Infections accelerate root system to rot.Root rot
To the yield effect of Radix Notoginseng it is huge be current notoginseng planting industry one of main bottleneck.
Since the prevention and treatment of current root rot is mainly by the method for a large amount of chemical pesticides of application, to medical material quanlity and safety
There is a degree of influence；Report related to the present invention is had no at present.
Summary of the invention
The object of the present invention is to provide one kind to clone from Radix Notoginseng and obtains PR genePnPR3, nucleotide
For sequence as shown in SEQ ID NO:1, which is 918bp, encodes the amino acid sequence as shown in SEQ ID NO:2
The protein of column.
Radix Notoginseng pathogenesis-related proteins PnPR3 of the present invention is also possible to the amino acid residue sequence in sequence table by one
Or substitution and/or deletion and/or addition and the protein relevant to plant disease-resistant of several amino acid residues.
The present invention is improving disease resistance of plant another object is that the gene of above-mentioned Radix Notoginseng pathogenesis-related proteins PnPR3 is applied
In.
Radix Notoginseng pathogenesis-related proteins PnPR3 gene of the present invention can be building up to existing prokaryotic expression carrier with existing method
It, can be before its transcription initiation nucleotide plus including constitutive promoter, enhanced promoter, induction or in plant expression vector
Any promoter including type promoter, tissue-specific promoter, stage of development specific promoter, for the ease of right
Turn PnPR3 gene cell or plant is identified and screened, used carrier can be processed, be such as added resistant
Antibiotic marker (ampicillin, gentamicin, kanamycins etc.) or that plant alternative label is added is (gus gene, glimmering
Light element enzyme gene etc.).The plant host that is converted is either monocotyledon, is also possible to dicotyledon, such as rice, small
Wheat, corn, cucumber, tobacco etc.；Carry PnPR3 gene of the invention expression vector can by application plant viral vector,
The conventional biology methods such as directly delivered DNA, mediated by agriculture bacillus convert plant cell or tissue, and by the plant of conversion through organizing
It is trained plant, obtains the plant that disease resistance improves.
The present invention expresses that it in tobacco by being transferred to foreign gene PnPR3 into tobacco, and experiment shows PnPR3
The overexpression of gene, to typical root rot germ Fusarium solani (Fusarium solani) there is certain inhibiting effect；This
The gene of invention can be transferred to the disease resistance that plant is improved in plant, and tobacco can be improved to root rot pathogenic bacteria Fusarium solani
(Fusarium solani) resistance, lay good working foundation, economy with higher for root rot Resistant breeding
Benefit.
Detailed description of the invention
Fig. 1 is the electrophoretogram that PnPR3 genetic fragment is expanded from the Radix Notoginseng main root that pine root fungus induces；
Fig. 2 is the prediction of PnPR3 albumen conserved domain；
Fig. 3 is PnPR3 phylogenetic tree analysis figure；
Fig. 4 is plant expression vector pK2GW7-35S-PnPR3Construct result verification；A figure isPnPR3The recycling of segment glue；In B figure
1. pENTRTM2B empty plasmid；2. Entry clone plasmids pENTRTM2B double digestion；C figure is pK2GW7-35S-PnPR3 Bacterium solution
PCR detection；D figure is pK2GW7-35S-PnPR3 Plasmid extracts；
Fig. 5 is the PCR electrophoresis detection schematic diagram of Agrobacterium bacterium colony；
Fig. 6 is that PCR detects transgene tobacco screening figure；
Fig. 7 is the PR-PCR verifying of PnPR3 gene in genetically modified plants；1: wild-type tobacco genomic DNA PCR electrophoresis is (negative
Control)；2: water PCR electrophoresis (blank)；3:pMD-18TPnPR3 plasmid electrophoresis (positive control)；4 ~ 7: it is followed successively by No. 4 respectively, 6
Number, No. 9, No. 11 plant DNA PCR electrophoresis；
Fig. 8 is to be overexpressed PnPR3 transgene tobacco to the resistance figure of Fusarium solani；
Fig. 9 is the chitinase activity figure for being overexpressed PnPR3 transgene tobacco.
Specific embodiment
Test method in embodiment described below is conventional method unless otherwise specified, and agents useful for same is such as without special
Explanation is conventional commercial reagent and the reagent that configures according to a conventional method.
Embodiment 1: the acquisition of Radix Notoginseng pathogenesis-related proteins PnPR3 conservative fragments and its cDNA overall length
Transcript profile sequencing filters out the significant pathogenesis-related proteins PR3 of differential expression, and the splicing of PnPR3 is obtained from CDS file
Sequence analyzes the overall length PR3 sequence of this segment species ginseng similar in MEGA software and Radix Notoginseng, it is found that the segment has
There are 3 ' ends and 5 ' ends.Therefore, this experiment only need to carry out high-fidelity PCR amplification to cDNA sequence, so that it may obtain the overall length of the gene
Sequence；Design special primer PR3-F (5 '-GGATCCatgagattttggacagtaaccatattc-3 ', GGATCC BamHI
Restriction enzyme site) and PR3-R (5 '-CAGCTGtagacctaaacctaaaccattaaatatgg-3 ', GTCGAC be SalI digestion
Site), using the open reading frame for the High fidelity PCR reaction enzymatic amplification gene that Dalian treasured biotech firm provides, obtain
The band of 1000bp or so size is consistent with the size of target gene；DNA fragmentation is recycled from Ago-Gel, connects pMD-
18T cloning vector is transformed into E.coli DH5 α, carries out bacterium colony PCR detection, extracts plasmid, carries out double digestion detection, is determined
After capable of being cut open, plasmid is sent and holds up biology Co., Ltd, section to Kunming and is sequenced；The final open reading for determining PnPR3 gene
Frame simultaneously obtains the cDNA segment containing restriction enzyme site (as shown in SEQ ID NO:1).
By Radix Notoginseng PR genePnPR3ORF(Open Reading Frame in sequence inputting NCBI
Finder) (www.ncbi.nlm.nih.gov/gorf/gorf.html) and DNAMAN software analytical sequence, as the result is shown
PnPR3 full length gene 918bp encodes 305 amino acid (as shown in SEQ ID NO:1)；Meanwhile using ExPASY
(ProtParam) protein molecular weight of software analysis PnPR3 coding is 33393.25 dalton, and Theoretical pi 6.41 divides
Minor is C1471H2200N408O449S19；Contain 26 basic amino acids (R, H, K), 23 acidity in 305 amino acid of PnPR3
Amino acid (D, E), nonpolarity, hydrophobic amino acid (G, A, V, L, I, F, P) have 132, non-ionized polarity, neutral amino
Sour (W, S, T, C, Y, M, N, Q) has 124；Wherein negatively charged amino acid includes D and E, amounts to 23, positively charged ammonia
Base acid includes R, H and K, and totally 26.
Utilize online tool InterPro(http: //www.ebi.ac.uk/interpro/search/sequence-
Search) PnPR3 albumen prediction conserved domain is analyzed, discovery PnPR3 full length protein is a conserved domain man
Race is glycosyl hydrolase family 19(Glycoside hydrolase, family19 (IPR016283)).The family is considered only
Contain chitinase activity.A Chitin-binding is contained in the family, and 1 structure of type (IPR001002) is located at 21 ~ 57
Position, 1 Lysozyme-like domain(lysozyme) (IPR023346) structure, it is located at 66 ~ 305.There are four the albumen contains
Region has chitinase catalytic activity, 211 ~ 221,69 ~ 298,70 ~ 298,87 ~ 109 is located at, 31 ~ 50
Also contain conserved positions a Chitin-binding, type 1 in position.PnPR3 protein bulk (GO) prediction shows that the albumen is joined
With carbohydrate metabolism process (carbohydrate metabolic process), chitin catalytic process (chitin
Catabolic process) and cell wall bulky molecular catalysis process (cell wall macromolecule catabolic
The biological processes such as process)；There is the albumen chitinase activity (chitinase activity) and chitin binding to live
The molecular functions such as property (chitin binding).
Carry out homology analysis using DNAMAN software, by Radix Notoginseng PnPR3 albumen and ginseng, carrot, walnut, spinach,
The albumen of cocoa, pigeonpea etc. constructs phylogenetic tree, sees Fig. 3；Radix Notoginseng PnPR3 albumen and ginseng PR3 albumen (accession number:
ACN96317.1) gather for one kind, show closer affiliation.
Experimental example 2: Radix Notoginseng PR genePnPR3Plant expression vector pK2GW7The building of -35S-PnPR3
Sequencing is detected into correct pMD-18T-PnPR3 vector plasmid restriction enzyme BamHI and SalI double digestion, recycling
PnPR3 genetic fragment；By pENTRTM2B plasmid carries out double digestion with restriction enzyme BamHI and SalI, recycles linear carrier
Segment；The PnPR3 genetic fragment of recycling and linear plasmid carrier after the recovery are attached, pENTR- can be successfully constructed
PnPR3 entry vector；The introduction expression vector is subjected to digestion detection according to Gateway kit operating instruction, will be built
Entry vector and plant expression vector pK2GW7.0 carries out LR reaction, can successfully construct pK2GW7The expression of -35S-PnPR3 plant
Carrier；The plant expression vector is subjected to PCR detection, digestion detection, plasmid is sequenced, and verifying the gene, there is no mutation
Carry out next step experiment (Fig. 4) afterwards.
Experimental example 3: Agrobacterium is converted with pK2GW7-35S-PnPR3 plant expression vector
Agrobacterium competent cell is prepared, with electric shock conversion method by the above-mentioned plant expression vector pK2GW7-35S- built
PnPR3 is transferred in Agrobacterium (C58C1 (pPMP90)), screens transformant on the LB plate added with spectinomycin.Take a small amount of matter
Grain is added in Agrobacterium competent cell, mixes gently, adds mixture in the electrotransformation cup of pre-cooling, gently tap cup body
So that mixed liquor is dropped down onto bottom of a cup, electrotransformation cup is placed in electric converter (BIO-RAD) sliding slot, click cup and 200 Europe with 1 mm
The parameter of nurse, 2.5 Kv/0.2cm shocks by electricity, and takes out electrotransformation cup after click immediately, is rapidly added 0.5mL LB culture medium,
It mixes, is transferred in the centrifuge tube of 1.5 mL；28 DEG C, 200 rpm shaking table culture 3-5 h；At room temperature, 7500 rpm centrifugation 1
Min abandons most of supernatant, retains 50 μ l and cell is resuspended；By the Agrobacterium after resuspension be coated on added with spectinomycin (Spe,
50 μ g/mL) LB solid medium on, 28 DEG C of cultures, 2 days acquisition single colonies, picking single colonie is in 20mL added with spectinomycin
On the LB liquid medium of (Spe, 50 μ g/mL), after 28 DEG C, 180rmp shaking table culture 36 hours, above and below PnPR3 gene
Trip primer makees PCR detection, and the band (Fig. 5) of 1.0kp, the transformant confirmed through bacterium colony PCR can be amplified by converting successful bacterium colony
Bacterium colony is for converting plant.
Experimental example 4: the Genetic Transformation in Higher Plants of mediated by agriculture bacillus and the screening of genetically modified plants
Receptor tobacco seed is seeded on 1/2 MS solid medium (pH5.8) after NaClO surface sterilization, is placed in 25 DEG C
The tissue culture room of constant temperature and continuous light (100 μm of ol/m2s) is cultivated 4 weeks, is obtained aseptic seedling, is cultivated in tissue culture room.Choose reason
Think that the tobacco leaf of state as explant, disseminates 20min in the Agrobacterium bacterium solution prepared, blots surface moisture, by leaf
Piece, which is laid on MS1 culture medium, dark co-cultures the time required when 2d(visually sees thallus with first blade), by tobacco
Blade explant is transferred on the bud inducement cultivation base such as MS4 of the factor containing screening and is screened (face down), about 15d subculture
Once.Blade edge can be observed in 20 ~ 25 days in transformed tobacco leaf explant, the illumination cultivation in subculture medium
Callus or Multiple Buds are grown, root media MS(is transferred to and contains antibiotic Cef+Km), the root induction under 25 DEG C of illumination.
It selects containing resistant transgene tobacco.
The genome of tobacco is extracted using CTAB method: being weighed 0.1 g plant leaf blade and is placed in 1.5mL centrifuge tube, liquid feeding nitrogen
It is ground to powdery.2 × CTAB buffer (Tris-HCl pH7.5 100 mM, EDTA 20 that 900 μ l are preheating to 65 DEG C is added
MM, NaCl 1.4 M, CTAB 2%), 65 DEG C of 20 min of water-bath, centre shakes up once every 2 min, and taking-up is cooled to room temperature, then
500 μ l chloroforms are added: isoamyl alcohol (24:1) mixed liquor, rotation shake up, and 4 DEG C, 7500 rpm, are centrifuged 10 min, shift supernatant
The centrifuge tube new to one, repeats the above steps；1/10 volume, 3 M pH5.2 NaOAc and isometric isopropanol is added, shakes up
4 DEG C afterwards, 12000 rpm, be centrifuged 20 min, abandon supernatant, cleaned with 75% ethyl alcohol and dried afterwards twice, with 1 containing RNase ×
TE buffer solution and degradation of rna, obtain the genomic DNA of transgene tobacco, using the DNA as template, with the spy of PnPR3 gene
Specific primer carries out PCR amplification, in doubtful PnPR3 transgenic plant, detects 4 after antibiotic-screening at 12 plants of acquisition
Strain genetically modified plants (T4, T6, T9, T11), PCR product band is single, and size meets PnPR3 sequence fragment length (Fig. 6).Through
The transgenic plant of PCR confirmation is analyzed for RT-PCR.
In order to investigate the transcription situation of the PnPR3 gene in the transgene tobacco strain containing gene, from genetically modified plants
Middle extraction total serum IgE, reverse transcription detect the transcription water of AtKUP1 gene in transgenic plants at RT-PCR analysis is used for after cDNA
It is flat.Using TRIzoL Reagent(RNA extract reagent, Invitrogen) extract RNA after, use RevertAidTM-M μ lV
Reverse Transcriptase Kit(reverse transcription reagent box, Fermentas) carry out cDNA synthesis, plant total serum IgE is about
0.1-0.5 ug, Oligo(dT) 50 ng, 10 mMdNTP mix, 1 μ l, 9 μ l are complemented to DEPC processing water, it is of short duration after mixing
It is collected in tube bottom by centrifugation, is placed in 65 DEG C of heating 5 min, 10 min of ice bath, and addition 11 μ l(5 of reaction mixture ×
Reaction buffer 4 μ l, 25 mM MgCl2 4 μ l, 0.1 M DTT2 μ l, 1 μ l of RNase inhibitor), by above-mentioned mixing
Object mixes, it is collected in tube bottom by of short duration centrifugation, and 25 DEG C of 2 min of heat preservation, then 42 DEG C of 70 min of heat preservation, synthesize cDNA.With
CDNA is template, makees PCR with PnPR3 gene upstream and downstream specific primer, the plant for being successfully transferred to PnPR3 can amplify 1.0
The band (Fig. 7) of kb, it was demonstrated that PnPR3 gene can be transcribed successfully in transgenic plants.The transgenosis confirmed through RT-PCR
Plant is used for further physiological and biochemical analysis.
Experimental example 4: overexpression PnPR3 gene pairs Fusarium solani (Fusarium solani) resistant proof.
In order to evaluate the resistance level of PnPR3 transgene tobacco, the tobacco lines chosen successfully for being transferred to PnPR3 gene
(T4, T6, T9, T11) and the healthy spire for cutting off transgene tobacco and control group.Evenly sized wound is formed on blade,
With the F. solani spore suspension of 200 μ L of sterile pipette tips inoculation, (spore concentration is 1 × 105Conidium/ml).It will place
The leaf managed is placed on sterile and wet filter paper, and is co-cultured in glass dish at 28 DEG C, and the morbidity feelings of leaf are observed
Condition.After 7 days, there is serious yellow and slightly rots in control tobacco leaf, and the blade of transgenic tobacco line is only shown slightly
Yellow；Blade is collected, by praising the chitinase kit that biological Co., Ltd buys from Yunnan scheme, uses spectrophotometry
Measure the chitinase activity in blade.Overexpression of the PnPR3 in tobacco can be by improving chitinase activity enhancing
Transgene tobacco pairF. solani Resistance (Fig. 8)；
Analysis inoculationF.solaniChitinase activity in the transgene tobacco and control blade of fungal spore；In initial rank
Section, the chitinase activity level in Tobacco Leaf maintain low-level, and transgene tobacco leaf due to PnPR3 overexpression and obtain
It obtains than compareing higher chitinase activity.With F. solani After spore inoculating 2 days, chitinase activity in tobacco leaf
Significant increase, chitinase activity is accumulated at any time in blade.Correlative study shows that higher plant is typically free of chitin, but
When by bacterium, fungi or virus infection, plant chitinase activity increases sharply, and enhancing plant resists pathogenic microorganism
Property；Therefore, PnPR3 is the important defensin gene (Fig. 9) for participating in infecting the anti-root rot of Radix Notoginseng.
Sequence table
<110>Kunming University of Science and Technology
<120>gene of Radix Notoginseng pathogenesis-related proteins PnPR3 and its application
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 918
<212> DNA
<213>Radix Notoginseng (Panax notoginseng)
<400> 1
atgagatttt ggacagtaac catattcatt ctagcctcat accttgcagt ttctgcagaa 60
caatgtggga aacaagctgg aatggctttg tgcccaaatg ggctctgttg cagccaattc 120
gggtggtgtg gcagcacccc tgagtactgc actaattgcc aaagccagtg cagcgaacct 180
tccccaggtg gaggtgttag ctccataatt actgagtctg tgtttaatca aatgcttaaa 240
tatcgaaacg acggaaggtg ccgtactaat ggattctaca catacaatgc ttttatcaat 300
gctgcaaaat ctttcaatgg ttttgggaca actggtaata ctgtccaaca gaaacaagag 360
ctcgccgctt tcttggctca gacctctcat gaaaccacag gtggatgggc aagtgcccca 420
gatggtcaat atgcatgggg atattgcttt ataagagaaa acaaccaggc tgcttactgc 480
acttccaaag actggccttg tcctcctggc aaactatact atggcagagg acctatccaa 540
ctcactcaca actacaacta tgggcaagct ggaaatgcaa ttggagtgga cctaataaac 600
aaccctgacc tagttgccac ggacgctacc atatcattca aaacagccat atggttctgg 660
atgactccac aggctaacaa gccatcgagc cacgatgtga ttacacaaag atggtcaccc 720
tctgcagcag ataccgcggc tggtcgcgtc cctggcttcg gtgtcatcac taacataatt 780
aacggcgggc tcgagtgtga tcacggcagg gatgataagg cggaggatag gattggattc 840
tacaagaggt attgtgacat tctgcaagtt ggctatggga atgaactcaa ttgcaacaat 900
caaaggcctt ttggctaa 918
<210> 2
<211> 305
<212> PRT
<213>Radix Notoginseng (Panax notoginseng)
<400> 2
Met Arg Phe Trp Thr Val Thr Ile Phe Ile Leu Ala Ser Tyr Leu Ala
1 5 10 15
Val Ser Ala Glu Gln Cys Gly Lys Gln Ala Gly Met Ala Leu Cys Pro
20 25 30
Asn Gly Leu Cys Cys Ser Gln Phe Gly Trp Cys Gly Ser Thr Pro Glu
35 40 45
Tyr Cys Thr Asn Cys Gln Ser Gln Cys Ser Glu Pro Ser Pro Gly Gly
50 55 60
Gly Val Ser Ser Ile Ile Thr Glu Ser Val Phe Asn Gln Met Leu Lys
65 70 75 80
Tyr Arg Asn Asp Gly Arg Cys Arg Thr Asn Gly Phe Tyr Thr Tyr Asn
85 90 95
Ala Phe Ile Asn Ala Ala Lys Ser Phe Asn Gly Phe Gly Thr Thr Gly
100 105 110
Asn Thr Val Gln Gln Lys Gln Glu Leu Ala Ala Phe Leu Ala Gln Thr
115 120 125
Ser His Glu Thr Thr Gly Gly Trp Ala Ser Ala Pro Asp Gly Gln Tyr
130 135 140
Ala Trp Gly Tyr Cys Phe Ile Arg Glu Asn Asn Gln Ala Ala Tyr Cys
145 150 155 160
Thr Ser Lys Asp Trp Pro Cys Pro Pro Gly Lys Leu Tyr Tyr Gly Arg
165 170 175
Gly Pro Ile Gln Leu Thr His Asn Tyr Asn Tyr Gly Gln Ala Gly Asn
180 185 190
Ala Ile Gly Val Asp Leu Ile Asn Asn Pro Asp Leu Val Ala Thr Asp
195 200 205
Ala Thr Ile Ser Phe Lys Thr Ala Ile Trp Phe Trp Met Thr Pro Gln
210 215 220
Ala Asn Lys Pro Ser Ser His Asp Val Ile Thr Gln Arg Trp Ser Pro
225 230 235 240
Ser Ala Ala Asp Thr Ala Ala Gly Arg Val Pro Gly Phe Gly Val Ile
245 250 255
Thr Asn Ile Ile Asn Gly Gly Leu Glu Cys Asp His Gly Arg Asp Asp
260 265 270
Lys Ala Glu Asp Arg Ile Gly Phe Tyr Lys Arg Tyr Cys Asp Ile Leu
275 280 285
Gln Val Gly Tyr Gly Asn Glu Leu Asn Cys Asn Asn Gln Arg Pro Phe
290 295 300
Gly
305
<210> 3
<211> 33
<212> DNA
<213>artificial sequence (Artificial)
<400> 3
ggatccatga gattttggac agtaaccata ttc 33
<210> 4
<211> 35
<212> DNA
<213>artificial sequence (Artificial)
<400> 4
cagctgtaga cctaaaccta aaccattaaa tatgg 35
Claims (2)
1. a kind of gene of Radix Notoginseng pathogenesis-related proteins PnPR3, nucleotide sequence is as shown in SEQ ID NO:1, coding such as SEQ
The protein of amino acid sequence shown in ID NO:2.
2. the gene of Radix Notoginseng pathogenesis-related proteins PnPR3 described in claim 1 is improving the application in disease resistance of plant.
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Cited By (2)
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|---|---|---|---|---|
| CN113322257A (en) * | 2021-05-31 | 2021-08-31 | 昆明理工大学 | Panax notoginseng inducible promoter PPO1 and application thereof |
| CN113652426A (en) * | 2021-08-20 | 2021-11-16 | 昆明理工大学 | Panax notoginseng inducible promoter R1 and application thereof |
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| CN113652426A (en) * | 2021-08-20 | 2021-11-16 | 昆明理工大学 | Panax notoginseng inducible promoter R1 and application thereof |
| CN113652426B (en) * | 2021-08-20 | 2023-06-20 | 昆明理工大学 | Pseudo-ginseng inducible promoter R1 and application thereof |
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