CN108866056A - Sweet potato IbCBF3 gene abiotic stress specific expressed promoter and its application - Google Patents
Sweet potato IbCBF3 gene abiotic stress specific expressed promoter and its application Download PDFInfo
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- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
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Abstract
The present invention relates to a kind of promoter that abiotic stress is specific expressed more particularly to sweet potato IbCBF3 gene abiotic stress specific expressed promoters and its application, belong to plant genetic engineering field.The present invention provides a kind of promoter that sweet potato IbCBF3 abiotic stress is specific expressed, and the nucleotide sequence of the promoter is as shown in SEQ ID NO.1；DNA fragmentation of the invention is the promoter of key transcription factor IbCBF3 in sweet potato low temperature stress approach, which contains multiple adverse circumstances and hormone response element.By luciferase network analyses double in tobacco leaf, which is quickly combined with upstream gene IbICE1 under low temperature stress, induces the expression of downstream gene, can be used for the research of sweet potato lower temperature resistance and transgenic breeding.
Description
Technical field
The present invention relates to a kind of promoter that abiotic stress is specific expressed more particularly to the non-lifes of sweet potato IbCBF3 gene
Object coerces specific expressed promoter and its application, belongs to plant genetic engineering field.
Background technique
Temperature is one of most important environmental factor in plant growth and development process.Low temperature not only influences crop growth hair
It educates and yield, also restricts farming time and the geographical distribution of crops.According to Food and Agricultural Organization of the United Nations (FAO, Food and
Agriculture Organization of the United Nations) latest data statistics, China's sweet potato cultivated area
It is 3,370,000 hectares, accounts for the 40% of world's cultivated area, be the largest sweet potato planting state；7,130,000 tons of total output, account for world sweet potato
The 73% of yield.It is daily to be widely applied to people because of its impoverishment tolerant, manageability, the high and abundant nutritive value of yield for sweet potato
It is the fourth-largest grain product after rice, wheat and corn in consumption, animal feed and industrial fuel.However sweet potato
Happiness temperature can not resist cold, similar to rice, corn, cotton, vulnerable to low temperature effect, cause the underproduction.No matter sweet potato seedling growth or receipts
It is stored after obtaining all very sensitive to low temperature.When temperature drops to 15 DEG C, sweet potato just stops growing, and is lower than 9 DEG C, potato wedge will gradually be cooled
It does harm to and rots；Overground part cauline leaf after frost quickly devitalization and it is dead.The not lower temperature resistance of sweet potato seriously constrains sweet potato
The development of industry.
For various abiotic stresses such as reply low temperature, plant forms complicated regulated and control network to perceive, respond and adapt to
The variation of external environment.ICE-CBF transcription factor pathway is studied the most deep in Chillingresistance Mechanisms of Plants.In many plants,
The winter resistance of plant can be improved by being overexpressed ICE or CBF.In arabidopsis, CBF3 transcription factor not only regulates and controls a large amount of downstreams
The expression of adversity gene, while the expression of its own is also by the effect of upstream gene.Its upstream transcription factor ICE1 can be tied
Closing on the MYC element (or ICE box) of CBF3 promoter activates the expression of CBF3 with response low temperature stress.
We draw according to the IbCBF3 gene order cloned in sweet potato main breed Xushen21 well, design in the present invention
Object obtains the promoter sequence of the gene by the method for chromosome walking.With the plant care software in PLACE to the sequence
Column are analyzed, and promoter element prediction result is shown：In IbCBF3 promoter in addition to transcribe necessary TATA-box,
GATA-box with Conserved Elements such as CAAT-box outside, there is also some induction environment-stress, light, moisture, hormone response and growths
The element of growth adjustment, such as response participate in ACE, ATCT-motif, GATT-motif, G-box and I-box that light is adjusted；It is de-
Fall sour response factors ABRE；Ethylene response factor ERE and methyl jasmonate response factors CGTCA-motif etc..Furthermore the starting
Son also comprising it is multiple can in conjunction with the regulatory factor ICE gene of upstream MYCRE cis-acting elements CANNTG etc..And utilize double fluorescence
Expression system, by the transient expression of tobacco leaf, verify the promoter can be incorporated into upstream gene under cryogenic, lure
Lead the expression for having correlation gene with low temperature stress.
Summary of the invention
The present invention is intended to provide promoter and its application that sweet potato IbCBF3 gene abiotic stress is specific expressed, this is opened
Mover belongs to the promoter of key transcription factor CBF3 in low temperature response approach, and content is related to its cDNA full length sequence, functional domain
Analysis can induce the expression of downstream gene in conjunction with upstream gene IbICE1 under gene low temperature stress induction.
The first purpose of the invention is to provide a kind of promoter that sweet potato IbCBF3 abiotic stress is specific expressed, institutes
The nucleotide sequence of promoter is stated as shown in SEQ ID NO.1.
The primer pair of any segment of sweet potato IbCBF3 promoter is expanded as shown in NO.2~11 SEQ ID.
The primer pair of the sweet potato IbCBF3 promoter overall length is expanded as shown in NO.12~13 SEQ ID.
Sweet potato IbCBF3 upstream gene IbICE1, it is characterised in that cDNA piece shown in SEQ ID NO.14 in sequence table
Section.
The primer pair of the sweet potato IbICE1 promoter overall length is expanded as shown in NO.15~16 SEQ ID.
The side of the specific expressed promoter of building sweet potato IbCBF3 gene abiotic stress is provided in the embodiment of the present invention
Method,
(1) it to the clone of sweet potato IbCBF3 gene promoter, using sweet potato cDNA as template, to the sequence PCR amplification, obtains
Sweet potato IbCBF3 gene promoter sequence overall length；
(2) sweet potato IbCBF3 upstream gene IbICE1 is constructed, is connected on T-blunt carrier, and converted to large intestine
In bacillus DH5 α competent cell, recombinant vector IbCBF3pro-GUS and CaMV35S-IbICE1 are obtained；
(3) transfer vector plasmid is converted respectively into agrobacterium strains GV3101, and single colonie base is extracted in YEP culture medium
Because of a group progress PCR verifying.
The second object of the present invention is to provide the recombinant vector containing the promoter.
In a kind of embodiment of the invention, the recombinant vector is IbCBF3pro-GUS and CaMV35S-IbICE1.
The present invention also provides a kind of buildings of the double luciferase expression systems of tobacco, contain recombinant vector, recombinant bacterium, transgenosis
Cell line or table expression cassette.
It is described to contain DNA fragmentation shown in above-mentioned SEQ ID NO.1 and SEQ ID NO.14.
The third object of the present invention is the heredity using the specific expressed promoter of above-mentioned sweet potato low temperature induction in plant
Application in breeding.
The promoter is the nucleic acid sequence of 2118 bit bases in nucleotide sequence, and the nucleic acid sequence encoding one is by low
Transcription factor temperature induction, that cold acclimation protein can be regulated and controled.
The promoter region includes the cis-acting elements such as 4 ABRE, 1 ERE and 1 CGTCA-motif, can be special
To hormone responses such as abscisic acid, ethylene and methyl jasmonates.The promoter also includes 5 MYCRE cis-acting elements
CANNTG, can be in conjunction with the regulatory factor IbICE1 gene of upstream.
Dual-luciferase reportor systerm carrier is constructed using the gene promoter and upstream regulating genes IbICE1, and is utilized
Mediated by agriculture bacillus technology injects tobacco leaf.Experiment shows that when tobacco is by low temperature stress, IbICE1 can be with the gene promoter
Sub strong combination, the expression of induced reporter gene (gus gene).
Beneficial effects of the present invention：
DNA fragmentation of the invention is the promoter of key transcription factor IbCBF3 in sweet potato low temperature stress approach, the starting
Son contains multiple adverse circumstances and hormone response element.By luciferase network analyses double in tobacco leaf, the promoter is in low temperature
Stress is lower quickly to be combined with upstream gene IbICE1, is induced the expression of downstream gene, be can be used for sweet potato lower temperature resistance and grind
Study carefully and transgenic breeding.
Detailed description of the invention
Fig. 1 is double fluorescent expression vector schematic diagrames
Fig. 2 is promoter in tobacco leaf by low temperature induction combination upstream gene
Specific embodiment
The clone of 1 sweet potato IbCBF3 gene promoter of embodiment
According to the cDNA sequence of transcription factor IbCBF3 important in sweet potato low temperature stress signal transduction path, press
The genome walker kit specification of Clontech company requires to set using the full length sequence of acquired IbCBF3 gene
Count 3 specific primers (SEQ ID NO.2:SP1:5'-GAAGTTGAGACAGGCGGAGCAG-3'；SEQ ID NO.3SP2:
5'-GTCTCCCGAAACTTCTTCCTCCC-3'；SEQ ID NO.4SP3:5'-CCAACAACACCTCTTCATCCGACA-3',
Design direction is the zone of ignorance direction for needing to expand, and the position of SP2 is located at the inside of SP1, and SP3 is located at the inside of SP2) and examination
The four kinds of annealing temperatures Jing Guo unique design provided in agent box lower degenerate primer AP1, AP2, AP3, AP4 are primer progress
Nested PCR amplification.It is sequenced after amplification is carried out glue recycling.Design NO.5~6 SEQ ID (SEQ ID NO.5:5'-
TGATAGTGAGTTGGGTTAGC-3；SEQ ID NO.6:5 '-TCCATATACGGCGGTACTT-3 '), using sweet potato cDNA as mould
Plate, to the sequence PCR amplification, amplified production is sequenced after carrying out glue recycling, verifies the sequence.And according to sequencing result, design the
Special primer (the SEQ ID NO.7SP4 of secondary genome walking:5'-GGAGTGTGATAAGGAGACAG-3'；SEQ ID
NO.8SP5:5'-CGCTACTGAAAGAGACAACA-3'；SEQ ID NO.9SP6:5'-CCTTCACGGTGCAGAATG-3',
The position of SP6 is located at the inside of SP5, and SP5 is located at the inside of SP4) and kit in provide four kinds moving back by unique designs
Fiery temperature lower degenerate primer AP1, AP2, AP3, AP4 are that primer carries out nested PCR amplification, and amplification is carried out glue recycling
After be sequenced.Design NO.10~11 SEQ ID (SEQ IDNO.10:5'-CATCTCGCCTAACTGTCAA-3'；SEQ ID
NO.11:5 '-GCTAACCCAACTCACTATCA-3 '), using sweet potato cDNA as template, to the sequence PCR amplification, amplified production into
It is sequenced after the recycling of row glue, verifies the sequence.Splice genome walking twice as a result, obtaining sweet potato IbCBF3 gene promoter
Sequence.
The sequence of 2 sweet potato IbCBF3 gene promoter of embodiment is analyzed
Plant care software in PLACE analyzes the sequence, as the result is shown：It is removed in IbCBF3 promoter
Outside the Conserved Elements such as transcription necessary TATA-box, GATA-box and CAAT-box, there is also some induction environment-stress,
The element that light, moisture, hormone response and growth and development are adjusted, such as response participate in ACE, ATCT-motif, GATT- that light is adjusted
Motif, G-box and I-box etc.；Abscisic acid response factors ABRE；Ethylene response factor ERE and methyl jasmonate response factors
CGTCA-motif etc..Furthermore the promoter also include it is multiple can in conjunction with the regulatory factor ICE gene of upstream MYCRE cis acting
Element CANNTG.
The building of 3 pairs of fluorescent expression vectors of embodiment
Using the cDNA of Xu's potato 29 as template, NO.12~13 primer SEQ ID (SEQ ID NO.125 '-is used respectively
GATTGGGAAGAAAGTAT-3'；SEQ ID NO.135 '-ATACGGCGGTACTTAGTAAAC-3 ') amplification IbCBF3 starting
Sub- full length sequence and NO.15~16 primer SEQ ID (SEQ ID NO.155 '-ATGTTATCAAGAGTGACAAGC '；SEQ ID
NO.165 '-GATCATCCCGTTGAAGCTAG ') amplification IbICE1 gene, be connected on T-blunt carrier, and converted to
In bacillus coli DH 5 alpha competent cell.Kanamycins is added in LB culture medium, through plate screening, obtains single colonie, extracts
Single colonie genome carries out PCR verifying, shakes bacterium extraction plasmid by the single colonie of verifying and is sequenced.By TAKARA company
In-fusion kit specification requires to arrive IbCBF3 promoter (abbreviation IbCBF3pro) and IbICE1 are gene constructed respectively
In carrier pBI101 and pCAMBIA1200, recombinant vector IbCBF3pro-GUS and CaM35S-IbICE1 are obtained.
Analysis of the promoter by low temperature induction combination upstream gene in 4 tobacco leaf of embodiment
Transfer vector plasmid IbCBF3pro-GUS and CaM35S-IbICE1 are converted respectively into agrobacterium strains GV3101,
Kanamycins or chloramphenicol are added in YEP culture medium, through plate screening, obtains single colonie, extract single colonie genome into
Row PCR verifying.Picking Agrobacterium single colonie is seeded in liquid YEP medium, and 30 DEG C are incubated overnight.100 μ l are taken to be incubated overnight liquid
Be transferred in the fresh YEP culture medium containing 100 μM of acetosyringones (As), 30 DEG C culture 4-6 hours, culture solution OD600
Reach 0.8-1.0, thalline were collected by centrifugation.With dip dyeing liquid for shell (50mM Mes-KOH, 50mM MgCl2It suspends again with 200 μM of As)
Thallus is adjusted to final concentration OD respectively600After=1.5, tobacco leaf is injected, and is placed on room temperature and restores 2 days.Tobacco is moved at 4 DEG C
Reason 3 hours, while designing solely according to group (25 DEG C are handled 3 hours), carry out GUS staining analysis.
Although the present invention has been described by way of example and in terms of the preferred embodiments, it is not intended to limit the invention, any to be familiar with this skill
The people of art can do various change and modification, therefore protection model of the invention without departing from the spirit and scope of the present invention
Enclosing subject to the definition of the claims.
Sequence table
<110>Xuzhou Agriculture Science Inst., Xuhuai area, Jiangsu (Xuzhou Sweetpotato Center)
<120>Sweet potato IbCBF3 gene abiotic stress specific expressed promoter and its application
<130> 2018.4.24
<160> 16
<170> SIPOSequenceListing 1.0
<210> 1
<211> 2118
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 1
tgtgtaggat cacagagcag tggtccgttc ttaggcatcc taaatgagca gtcagcctaa 60
cacaacatat ttggaaaata ccacaaatat taccacctat tatatctaac acatgaatat 120
atgaaaaata ttatatgtac tctcattttc tactctcaac ttttatggca tgtttggttg 180
gatggaaaat caattccatg gaaaagattt tccaaaaagt tgaggaaaat gatgaataat 240
tatgttgttt ggttggatgg aaaatgtttt ccatggaatt tgacttccca aaagaaagga 300
aaataaattc tttggttgag ataggtattt tgttttccaa agagattggg aagaaagtat 360
aagcccttgg actattttac cctcatcaaa attgactaat tacacatgta tatatagcca 420
aaccattaat tactaaaggc atattgatct ttatattcat aattccttac cattccaaac 480
ccaaccaaat aatggaattc atatttccaa taatttcttt tccaccaacc aaacagtggg 540
atctattttc ctggtaccac ggaatttgaa tttcatttcc ttcaaatatt ttctagcaac 600
caaacgaact gttacttttc cacacaaatc ttgatataga tacttatatt gaaatctaca 660
tgaaaaaata gcttattagt gtccgtcata tcaagaaata aaatgtccat gtagtagata 720
tattggtaat aattgattct agaaaaagta atgtagtatt gtcacaaatt tggtacaata 780
cgacaagttg atactgagat ttgtgaatct ccagccttcc gtacaggtgt actaggtggt 840
agtaggtacc taggcgatga aagggacaaa ggagtagaga tttggtggtt ccttgacaag 900
atttgggcgg ggcatgggat gaagccatta ggcatgtggc tcacgcgttt ccgtggaaaa 960
accaccggcg ttttactgct tccaaccaat catcaatcct aataatgttt tttttttttt 1020
tgagaaatca atcctaataa tgttggcggt tgtaaaacta acatctcgcc taactgtcaa 1080
ccgcgtctat catctccatc tttcatcctc caattaactg tcacctcgct ccaaacaatc 1140
cttattctgt cctttcttcc atccatccat ctccaatcct taatctatac tatatgtttt 1200
ctgtactcta tgtattgctg attatatatt tattttaaag aaaattaata taaaatttga 1260
aattttacgt agccgtataa aaatcttgtg tgagaccgtt ttacgaatct taatctattt 1320
tggtttgtaa taatcttcag ataataaata tagttctcaa caagaattac atcaaaataa 1380
aaaatagtag gtcttgtatt aggacaatat aatcctcaaa tcttattttt taaaaaatag 1440
tattagtaag tataaaaagg tatttctaga aaataatatt tttattaaat tatatcaaac 1500
tagggaatta aaattaaatt gatagtgaaa ctgaagaggg actgagtata ctgtaacgtg 1560
tgtgcatggc ggaaaagtca tagtatctta ctcgtagtta tggaaatatg aagtacggaa 1620
gatattatga tagtgagttg ggttagctga actttgactt gatatattca gcattctgca 1680
ccgtgaaggt ccgtggaatc ccccgtaaac aagggggcgg gccccattca tgaagtgaaa 1740
gcacacgtgt cgggtacaca acttacgata ccttttaatt tcatgttgtc tctttcagta 1800
gcgtgaccca aatccagttg gcagtttgaa tgaccccacc acgtacagct ggctattcac 1860
ttcatcatat tttccatatt tacctcaatt cccagttcct tcctcctagg ctgtctcctt 1920
atcacactcc gtgttttttc gcgtctccaa cgtaagctta tcaccttaca cttatcgaac 1980
actccccccc tcacccaccc atacgcatac atatatatat aactcccatt ttaggtagtg 2040
tgttaattac ttcatacata cacactccat tttagctttt ccatatatat atatatactt 2100
tactaagtac cgccgtat 2118
<210> 2
<211> 22
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 2
gaagttgaga caggcggagc ag 22
<210> 3
<211> 23
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 3
gtctcccgaa acttcttcct ccc 23
<210> 4
<211> 24
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 4
ccaacaacac ctcttcatcc gaca 24
<210> 5
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 5
tgatagtgag ttgggttagc 20
<210> 6
<211> 19
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 6
tccatatacg gcggtactt 19
<210> 7
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 7
ggagtgtgat aaggagacag 20
<210> 8
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 8
cgctactgaa agagacaaca 20
<210> 9
<211> 18
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 9
ccttcacggt gcagaatg 18
<210> 10
<211> 19
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 10
catctcgcct aactgtcaa 19
<210> 11
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 11
gctaacccaa ctcactatca 20
<210> 12
<211> 17
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 12
gattgggaag aaagtat 17
<210> 13
<211> 21
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 13
atacggcggt acttagtaaa c 21
<210> 14
<211> 1638
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 14
atgttatcaa gagtgacaag catggtttgg atggatggga aagaggaaga acaagcaggt 60
tcttgggtac agaacaacaa tggcggcggt ggggccggag gaggaggatt agcgggcaag 120
gaggaaatgg agatggcaac aatcaagtcc atgctggaag ctgaggaagt ggagtggtac 180
atggctaata atcagcatag ccacaacaat ggtgcaccca tgcaaggcca tgggggcatt 240
tctttctcta caaatttctc tgagcctgac aacaatctga tcttgcaccc tgtggattcc 300
tcttcctctt gctctccttc ctctgcttct gttttcaatg ctcttgaccc ttctcaggtt 360
cactattttt tgccccacaa ggctgccatg atgagccatc ccttggatca gggtgggttt 420
gatttggggt gtgagagtgg gtttcttgaa actcaagccc tgagtggttt gagtagaggg 480
ggaggggttt tgggtggtgg gtttggtgat ttgagctgtc agaacttctt gggggctccc 540
aacttgagct ctgttcctca atttggttca acccatttgc tgcaacttcc acacaatggt 600
ggaggagggg ggtttggtcc actagggttt ggagagggct atgtgaatgt gaatgagaat 660
gagaatgaga atgctttgtt tcttaatagg tccaagttgt taaagccact tgataatttt 720
gcttcaattg gggcacagcc tactctcttt caaaagaggg ctgctcttag gaagaatctt 780
ggcaattcta gtggaaattt agcacttttg ggtggtgaaa ttggccacac tgatagcagc 840
tttgataaga agagtgaagt gaatgagagg aagaggaaag ggagcaatgg gggggatgaa 900
ttggaggatg tgagcattga tggctccaac ttgaactatg actcagatga gcttgtcgaa 960
aacagtggca aagttgatga aagtgtgaag aatggtggaa ttagctcctc caatgccact 1020
gggggtgacc aaaaggggaa gaagaaaggg cttccagcca agaacttgat ggccgaaagg 1080
aggcgtagga agaagctcaa cgacaggctt tacatgttga ggtctgttgt cccgaagatt 1140
agtaagatgg acagagcttc gattttaggg gatgcaattg aatacttgaa ggaacttctg 1200
cagaaaatca atgacctcca caatgaactc gagtctactc ctccttgctc cgcattaacc 1260
cctaattcga gtttctaccc gttgacacca actgcatctg ccctaccctg ccgtatcaaa 1320
gaagaaatca gtccaactgc atttgcaagc ccgctgtcta gtccaactgg acagcctgca 1380
agggttgaag taagggttag agaaggaaga gcggtgaata tccatatgtt ttgtagccgc 1440
aaacccggcc tattactttc aacaatgaag gctcttgaca accttggtat agacatccaa 1500
caggctgtta tcagctgctt caacgggttt gccttggata ttttccgagc agagcaatgc 1560
aaggaaggcc aagacatcca tccagatcaa atcaaagctg tacttatgga ttccgctagc 1620
ttcaacggga tgatctaa 1638
<210> 15
<211> 21
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 15
atgttatcaa gagtgacaag c 21
<210> 16
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 16
gatcatcccg ttgaagctag 20
Claims (10)
1. the specific expressed promoter of sweet potato IbCBF3 gene abiotic stress, it is characterised in that：The nucleosides of the promoter
Acid sequence is as shown in SEQ ID NO.1.
2. the specific expressed promoter of sweet potato IbCBF3 gene abiotic stress according to claim 1；Its feature exists
In：The primer pair of any segment of sweet potato IbCBF3 promoter is expanded as shown in NO.2~11 SEQ ID.
3. the specific expressed promoter of sweet potato IbCBF3 gene abiotic stress according to claim 2；Its feature exists
In：The primer pair of the sweet potato IbCBF3 promoter overall length is expanded as shown in NO.12~13 SEQ ID.
4. the specific expressed promoter of sweet potato IbCBF3 gene abiotic stress according to claim 3；Its feature exists
In：Construct sweet potato IbCBF3 upstream gene IbICE1, it is characterised in that cDNA segment shown in SEQ ID NO.14 in sequence table.
5. the specific expressed promoter of sweet potato IbCBF3 gene abiotic stress according to claim 4；Its feature exists
In：The primer pair of the sweet potato IbIDE1 promoter overall length is expanded as shown in NO.15~16 SEQ ID.
6. the specific expressed promoter of sweet potato IbCBF3 gene abiotic stress according to claim 5；Its feature exists
In：The method for constructing the specific expressed promoter of sweet potato IbCBF3 gene abiotic stress,
(1) to the clone of sweet potato IbCBF3 gene promoter, using sweet potato cDNA as template, to the sequence PCR amplification, sweet potato is obtained
IbCBF3 gene promoter sequence overall length；
(2) sweet potato IbCBF3 upstream gene IbICE1 is constructed, is connected on T-blunt carrier, and converted to Escherichia coli
In DH5 α competent cell, recombinant vector IbCBF3 pro-GUS and CaMV35S-IbICE1 is obtained；
(3) transfer vector plasmid is converted respectively into agrobacterium strains GV3101, and single colonie genome is extracted in YEP culture medium
Carry out PCR verifying.
7. containing any promoter of claim 1-6 and its upstream gene recombinant vector, recombinant bacterium, transgenic cell line or
Expression cassette.
8. promoter according to claim 7 and its upstream gene contain above-mentioned SEQ ID NO.1 and SEQ ID NO.14
Shown in DNA fragmentation.
9. the specific expressed promoter of sweet potato IbCBF3 gene abiotic stress described in claim 1 is educated in the heredity of plant
Application in kind.
10. promoter according to claim 9, it is characterised in that：For the nucleic acid of 2118 bit bases in nucleotide sequence
Sequence, the nucleic acid sequence encoding one by low temperature induction, the transcription factor of cold acclimation protein can be regulated and controled；The promoter region
Comprising cis-acting elements such as 4 ABRE, 1 ERE and 1 CGTCA-motif, can specifically to abscisic acid (ABA), ethylene and
The hormone responses such as methyl jasmonate (MeJA).The promoter also includes 5 MYCRE cis-acting elements CANNTG, energy and upstream
Regulatory factor ICE (INDUCER OF CBF EXPRESSION) gene combines.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114408898A (en) * | 2022-01-24 | 2022-04-29 | 江苏徐淮地区徐州农业科学研究所(江苏徐州甘薯研究中心) | Biomass carbon dots and preparation method and application thereof |
| CN114958876A (en) * | 2022-06-29 | 2022-08-30 | 河南省农业科学院植物营养与资源环境研究所 | Application of IAA-PO1 gene in inducing formation of oyster mushroom primordium and stress resistance of oyster mushroom growth and development |
-
2018
- 2018-07-05 CN CN201810730214.2A patent/CN108866056B/en active Active
Non-Patent Citations (1)
| Title |
|---|
| RONG JIN等: "Overexpressing IbCBF3 increases low temperature and drought stress tolerance in transgenic sweetpotato", 《PLANT PHYSIOLOGY AND BIOCHEMISTRY》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114408898A (en) * | 2022-01-24 | 2022-04-29 | 江苏徐淮地区徐州农业科学研究所(江苏徐州甘薯研究中心) | Biomass carbon dots and preparation method and application thereof |
| CN114408898B (en) * | 2022-01-24 | 2024-03-22 | 江苏徐淮地区徐州农业科学研究所(江苏徐州甘薯研究中心) | Biomass carbon dot and preparation method and application thereof |
| CN114958876A (en) * | 2022-06-29 | 2022-08-30 | 河南省农业科学院植物营养与资源环境研究所 | Application of IAA-PO1 gene in inducing formation of oyster mushroom primordium and stress resistance of oyster mushroom growth and development |
| CN114958876B (en) * | 2022-06-29 | 2023-07-21 | 河南省农业科学院植物营养与资源环境研究所 | Application of IAA-PO1 gene in inducing oyster mushroom primordium to form and oyster mushroom growth and development stress resistance |
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|---|---|
| CN108866056B (en) | 2021-05-18 |
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Application publication date: 20181123 Assignee: Xuzhou Youda Qingyuan Agricultural Technology Development Co.,Ltd. Assignor: JIANGSU XUHUAI DISTRICT XUZHOU AGRICULTURAL Research Institute (JIANGSU XUZHOU SWEET POTATO RESEARCH CENTER) Contract record no.: X2023980049637 Denomination of invention: Promoter of IbCBF3 gene in sweet potato and its application for non biological stress-specific expression Granted publication date: 20210518 License type: Common License Record date: 20231205 |