CN1243098C - Paddy rice anti-reverse transcripfactor and its coding gene and application - Google Patents
Paddy rice anti-reverse transcripfactor and its coding gene and application Download PDFInfo
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Abstract
The present invention discloses a stress resistant transcription factor of rice, a coded gene thereof and an application thereof. The aim of the present invention is to provide a transcription factor with the characteristic of strong stress resistance and a coded gene thereof. The stress resistant transcription factor of the present invention, which is called OsDREB1, is a protein with an amino acid residue sequence of sequence 2 in a sequence list, or a protein which is used for substituting, deleting or adding one or a plurality of amino acid residues of the amino acid residue sequence of sequence 2, has the same activity with the amino acid residue sequence of sequence 2, and is derived from sequence 2. The coded gene of the stress resistant transcription factor OsDREB1 is one of the following nucleotide sequences: 1) a DNA sequence of sequence 1 in the sequence list, and 2) a DNA sequence which has more than 90 % of homology with a DNA sequence limited by sequence 1 in the sequence list and codes the same functional protein. The gene of the present invention has important meanings for breeding stress resistant plant varieties, especially cold resistant plant varieties, and increasing crop yield.
Description
Technical field
The present invention relates to plant transcription factor and encoding gene thereof and application, particularly derive from transcription factor and encoding gene and the application of paddy rice.
Background technology
The variation of physical chemical factor in the environment, for example arid, saline and alkaline, damage to plants caused by sudden drop in temperature, the factor of coercing such as freeze injury, waterlogging has material impact to growth and development of plant, can cause the extensive underproduction of farm crop when serious, cultivating the resistance of reverse crop is one of major objective of plant husbandry.Improve the resistance of reverse of crop, except utilizing traditional breeding method, at present, one of field that the molecular genetic breeding has become the scientific worker to be paid close attention to.Higher plant cell has number of ways to experience the variation of physico-chemical parameter in the external environment, the signal that born of the same parents are outer becomes intracellular signal, through a series of phosphorylation level chain reactions signal is passed to transcription factor, transcription factor remakes and is used for functional gene, start the expression of adverse circumstance response gene, improve the resistance of reverse of plant.Verified, in plant, the class in the EREBP/AP2 class DREB transcription factor family can be accepted the environment-stress signal and start the adverse circumstance response gene.
Paddy rice is understood fully its anti-contrary mechanism, and then improves its resistance of reverse to have important theory and realistic meaning as most important food crop.
Summary of the invention
The purpose of this invention is to provide transcription factor and encoding gene thereof with strong adverse-resistant characteristic.
The anti-reverse transcription factor provided by the present invention derives from paddy rice, name is called OsDREB1, be protein, or the amino acid residue sequence of sequence 2 is passed through replacement, disappearance or the interpolation of one or several amino-acid residue and has identical active by sequence 2 deutero-protein with the amino acid residue sequence of sequence 2 with sequence 2 amino acid residue sequences in the sequence table.
The protein that sequence 2 amino acid residue sequences are made up of 238 amino-acid residues in the sequence table is a kind of DREB transcription factor in the paddy rice, contains conservative EREBP/AP2 structural domain sequence, in the sequence 2 from the nitrogen end to carbon teminal 51-117 amino acids residue.
The encoding gene of anti-reverse transcription factor OsDREB1 is the dna sequence dna of sequence 1 in the sequence table.
The dna sequence dna of sequence 1 is by 727 based compositions in the sequence table, and the reading frame of this gene is that it is expressed and mainly is subjected to cryogenic inducing from 3 ' end the 7th to the 723rd bit base.
Utilize any carrier that can guide foreign gene in plant, to express,, can obtain transgenic cell line and transfer-gen plant that the low temperature stress tolerance is enhanced the gene transfered plant cell of encoding transcription factor OsDREB1 provided by the present invention.Gene of the present invention can add any enhancing promotor or inducible promoter in being building up to plant expression vector the time before its transcription initiation Nucleotide.For the ease of transgenic plant cells or plant being identified and screening, can process employed carrier, as the antibiotic marker thing (gentamicin, kantlex etc.) that adds the alternative mark (gus gene, luciferase genes etc.) of plant or have resistance.Carry that OsDREB1 expression carrier of the present invention can Ti-plasmids, Ri plasmid, plant viral vector, directly DNA conversion, microinjection, electricity be led, conventional biological method transformed plant cells or tissue such as agriculture bacillus mediated by using, and the plant transformed tissue cultivating is become plant.By the plant transformed host both can be monocotyledons, also can be dicotyledons, as: paddy rice, wheat, corn, cucumber, tomato, willow, turfgrass, lucerne place etc.Gene pairs of the present invention is cultivated the adversity resistant plant kind, particularly cultivates the cold resistant plant kind, and it is significant to improve crop yield.
The present invention will be further described below in conjunction with drawings and the specific embodiments.
Description of drawings
Fig. 1 is the partial results of dot blot.
Fig. 2 is Northern hybridization analysis result.
Embodiment
The screening of embodiment 1, rice Os DREB1 transcription factor and the clone of cDNA thereof
80 amino acid conserved sequences with AP2 gene in the paddy rice carry out the BLAST retrieval in the paddy gene storehouse, obtain about 1000 reads fragments, and about 140 the reads fragment of wherein picking out is spliced with Phrap software, obtain 32 contig fragments.
Paddy rice (Oryza sativa var.JX17) seed kind is divided into two parts after one month in basin, a continued growth, and another part is carried out the low temperature stress processing, and the paddy rice seedling is moved into 4 ℃ of lighting box, sampling after 5 hours.Collect fresh blade 1g and grind in liquid nitrogen, be suspended from the 4mol/L sulphur hydracid guanidine, mixture adds dehydrated alcohol and precipitates total RNA with acid phenol, chloroform extracting in the supernatant, afterwards, water-soluble.Obtain mRNA through the processing of mRNA purification kit.Be probe and 32 the EREBP/AP2 genoid contig dot blots that screen of point on the Hybond-N+ film with above-mentioned mRNA normal and that coerce respectively.Secondary is hybridized discrepant gene select, carry out two again and take turns hybridization checking (result as shown in Figure 1, among the figure, first round dot blot: (A) low temperature was handled 5 hours for 4 ℃; (B) contrast, second takes turns dot blot: (C) low temperature was handled 5 hours for 4 ℃; (D) contrast.Underscore is illustrated in the inductive clone that all catches a cold in the two-wheeled dot blot.Arrow is represented the Actin gene as last sample internal reference.), obtain the dreb gene OsDREB1 of low temperature induction.
The relation of embodiment 2, rice Os DREB1 activity and environment-stress
Paddy rice (Oryza sativa var.JX17) seed kind is divided into two parts after one month in basin, a continued growth, and another part is carried out the following various processing of coercing:
4 ℃ of subzero treatment: the paddy rice seedling is moved into 4 ℃ of lighting box.
Salt is handled: the paddy rice seedling is moved in the 200mM NaCl solution.
ABA handles: the paddy rice seedling is moved in the 50uM ABA solution.
Drought is handled: blot with filter paper to be put in behind the moisture on paddy rice seedling surface it is dewatered naturally.
Respectively in the sampling in 0,0.5,1,2,5,10,24,32 hour of various processing back.Collect fresh blade 1g and grind in liquid nitrogen, be suspended from the 4mol/L sulphur hydracid guanidine, mixture adds dehydrated alcohol and precipitates total RNA with acid phenol, chloroform extracting in the supernatant, water-soluble afterwards, obtains total RNA.Be that probe carries out the Northern analysis with OsDREB1 DNA respectively.The result as shown in Figure 2, A is a subzero treatment among the figure; B is that 50umABA handles; C handles for 200mM NaCl; D is that arid is handled, and Northern analysis revealed, OsDREB are induced by low temperature (4 ℃), but are not subjected to 50uM ABA, 200mM NaCl and drought-induced.
Sequence table
<160>2
<210>1
<211>727
<212>DNA
＜213〉Oryza paddy rice (Orysa sativa L.)
<400>1
ccgaagatgt?gcgggatcaa?gcaggagatg?agcggcgagt?cgtcggggtc?gccgtgcagc 60
tcggcgtcgg?cggagcggca?gcaccagacg?gtgtggacgg?cgccgccgaa?gaggccggcg 120
gggcggacca?agttcaggga?gacgaggcac?ccggtgttcc?gcggcgtgcg?gcggaggggc 180
aatgccggga?ggtgggtgtg?cgaggtacgg?gtgcccgggc?ggcgcggctg?caggctctgg 240
ctcggcacgt?tcgacaccgc?cgagggcgcg?gcgcgcgcgc?acgacgccgc?catgctcgcc 300
atcaacgccg?gcggcggcgg?cggcggggga?gcatgctgcc?tcaacttcgc?cgactccgcg 360
tggctcctcg?ccgtgccgcg?ctcctaccgc?accctcgccg?acgtccgcca?cgccgtcgcc 420
gaggccgtcg?aggacttctt?ccggcgccgc?ctcgccgacg?acgcgctgtc?cgccacgtcg 480
tcgtcctcga?cgacgccgtc?caccccacgc?accgacgacg?aggaggagtc?cgccgccacc 540
gacggcgacg?agtcctcctc?cccggccagc?gacctggcgt?tcgaactgga?cgtcctgagt 600
gacatgggct?gggacctgta?ctacgcgagc?ttggcgcagg?ggatgctcat?ggagccacca 660
tcggcggcgc?tcggcgacga?cggtgacgcc?atcctcgccg?acgtcccact?ctggagctac 720
tagagct 727
<210>2
<211>238
<212>PRT
＜213〉Oryza paddy rice (Orysa sativa L.)
<400>2
Met?Cys?Gly?Ile?Lys?Gln?Glu?Met?Ser?Gly?Glu?Ser?Ser?Gly?Ser
l 5 10 15
Pro?Cys?Ser?Ser?Ala?Ser?Ala?Glu?Arg?Gln?His?Gln?Thr?Val?Trp
20 25 30
Thr?Ala?Pro?Pro?Lys?Arg?Pro?Ala?Gly?Arg?Thr?Lys?Phe?Arg?Glu
35 40 45
Thr?Arg?His?Pro?Val?Phe?Arg?Gly?Val?Arg?Arg?Arg?Gly?Asn?Ala
50 55 60
Gly?Arg?Trp?Val?Cys?Glu?Val?Arg?Val?Pro?Gly?Arg?Arg?Gly?Cys
65 70 75
Arg?Leu?Trp?Leu?Gly?Thr?Phe?Asp?Thr?Ala?Glu?Gly?Ala?Ala?Arg
80 85 90
Ala?His?Asp?Ala?Ala?Met?Leu?Ala?Ile?Asn?Ala?Gly?Gly?Gly?Gly
95 100 105
Gly?Gly?Gly?Ala?Cys?Cys?Leu?Asn?Phe?Ala?Asp?Ser?Ala?Trp?Leu
110 115 120
Leu?Ala?Val?Pro?Arg?Ser?Tyr?Arg?Thr?Leu?Ala?Asp?Val?Arg?His
125 130 135
Ala?Val?Ala?Glu?Ala?Val?Glu?Asp?Phe?Phe?Arg?Arg?Arg?Leu?Ala
140 145 150
Asp?Asp?Ala?Leu?Ser?Ala?Thr?Ser?Ser?Ser?Ser?Thr?Thr?Pro?Ser
155 160 165
Thr?Pro?Arg?Thr?Asp?Asp?Glu?Glu?Glu?Ser?Ala?Ala?Thr?Asp?Gly
170 175 180
Asp?Glu?Ser?Ser?Ser?Pro?Ala?Ser?Asp?Leu?Ala?Phe?Glu?Leu?Asp
185 190 195
Val?Leu?Ser?Asp?Met?Gly?Trp?Asp?Leu?Tyr?Tyr?Ala?Ser?Leu?Ala
200 205 210
Gln?Gly?Met?Leu?Met?Glu?Pro?Pro?Ser?Ala?Ala?Leu?Gly?Asp?Asp
215 220 225
Gly?Asp?Ala?Ile?Leu?Ala?Asp?Val?Pro?Leu?Trp?Ser?Tyr
230 235 238
Claims (7)
1, anti-reverse transcription factor OsDREB1, be protein, or the amino acid residue sequence of sequence 2 is passed through replacement, disappearance or the interpolation of one or several amino-acid residue and has identical active by sequence 2 deutero-protein with the amino acid residue sequence of sequence 2 with sequence 2 amino acid residue sequences in the sequence table.
2, transcription factor according to claim 1 is characterized in that: be the conservative territory of EREBP/AP2 function from the nitrogen end to carbon teminal 51-117 amino acids residue in the described sequence 2.
3, the encoding gene of anti-reverse transcription factor OsDREB1 is the dna sequence dna of sequence 1 in the sequence table.
4, gene according to claim 3 is characterized in that: the reading frame of this gene is for holding the 7th to the 723rd bit base from 3 '.
5, contain the described expression carrier of claim 3.
6, the transgenic cell line that contains the described gene of claim 3.
7, the application of the described gene of claim 3 in cultivating the plant with adverse resistance kind.
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| CN 02129517 CN1243098C (en) | 2002-08-29 | 2002-08-29 | Paddy rice anti-reverse transcripfactor and its coding gene and application |
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| CN 02129517 CN1243098C (en) | 2002-08-29 | 2002-08-29 | Paddy rice anti-reverse transcripfactor and its coding gene and application |
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| CN1478893A CN1478893A (en) | 2004-03-03 |
| CN1243098C true CN1243098C (en) | 2006-02-22 |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101062943B (en) * | 2007-04-29 | 2010-09-08 | 北京未名凯拓农业生物技术有限公司 | Rice stress tolerance related DREB transcription factor and its coding gene and application |
Families Citing this family (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100348723C (en) * | 2004-04-06 | 2007-11-14 | 北京未名凯拓农业生物技术有限公司 | Reverse-tolerant concerned gene of rice and its coding protein and use |
| CN1332980C (en) * | 2004-12-10 | 2007-08-22 | 中国科学院上海生命科学研究院 | Paddy rice anti contravariance related gene-anchor series repetitive protein gene and its application |
| CN1328288C (en) * | 2005-10-10 | 2007-07-25 | 中国科学院植物研究所 | Rice DREB transcription factor and its coding gene and application |
| CN1313486C (en) * | 2005-11-08 | 2007-05-02 | 北京北方杰士生物科技有限责任公司 | Plant DREB transcription factor and its coding gene and use |
| CN101037696B (en) * | 2006-03-16 | 2010-11-03 | 华中农业大学 | Paddy cool injury gene and application |
| CN100465190C (en) * | 2006-05-31 | 2009-03-04 | 北京凯拓迪恩生物技术研发中心有限责任公司 | Plant anti-reverse related protein, and its coding gene and use |
| CN101139385B (en) * | 2007-07-31 | 2010-06-09 | 中国科学院植物研究所 | Vegetable stress-resistant related protein and its coding gene and application |
| CN107663230A (en) * | 2016-07-28 | 2018-02-06 | 中国科学院植物研究所 | Application of the cold-resistant GAP-associated protein GAP in plant cold tolerance is regulated and controled |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101062943B (en) * | 2007-04-29 | 2010-09-08 | 北京未名凯拓农业生物技术有限公司 | Rice stress tolerance related DREB transcription factor and its coding gene and application |
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