CN103305488B - Plant drought resistance related protein as well as encoding gene and application thereof - Google Patents
Plant drought resistance related protein as well as encoding gene and application thereof Download PDFInfo
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Abstract
The invention discloses a plant drought resistance related protein, belonging to the field of plant gene engineering. The plant drought resistance related protein disclosed by the invention is composed of an amino acid sequence as shown by SEQ ID No: 1 and a derivative sequence thereof. The invention further discloses a gene of the protein, wherein the gene is composed of a nucleotide sequence as shown by SEQ ID No: 2 and a derivative sequence thereof. The invention further discloses usage of the gene for improving the plant drought resistance. The plant drought resistance related protein disclosed by the invention greatly increases the sucrose content of transgenic plants by over-expression of cy-FBPase and greatly improves the drought tolerance of the transgenic plants. The plant drought resistance related protein disclosed by the invention provides an effective way of improving the plant drought tolerance through genetic engineering.
Description
Technical field
The invention belongs to plant genetic engineering field, be specifically related to a kind of albumen relevant to drought resistance in plants, and the encoding gene of this albumen, also relate to this gene in the purposes improving in plant biomass and drought tolerance.
Background technology
In vegetable cell, there is fructose-1 of two types, 6-diphosphatase, it is respectively chloroplast(id) fructose-1,6-diphosphatase (referred to as: cp-FBPase) and tenuigenin fructose-1,6-diphosphatase (referred to as: cy-FBPase), the structural similitude of two enzymes, effect substrate is identical, all that the hydrolysis of catalysis fructose-1，6-diphosphatase becomes fructose-6-phosphate and inorganic phosphate.But the active position difference of two kinds of enzymes, enzymic activity regulative mode and action pathway are also different.Wherein cp-FBPase participates in Calvin cycle, is the enzyme that is subject to light regulation and control; Cy-FBPase activity is not regulated and controled by light.Between the cy-FBPase of different plants, can reach 80% homology, between the cp-FBPase of different plants, also can reach the homology of 80-90%, but between the FBPase of two types, homology but only has 50%, between them, there is not immunological cross-reaction, but but have similar albumen tertiary structure.Exist with single aggressiveness, dimer and tetrameric form in vivo, only have the tetramer just to there is catalytic activity.
Cy-FBPase does not belong to Calvin cycle, it is the key enzyme in photosynthetic organs sucrose route of synthesis, also be the reference mark that in glyconeogenesis, regulation and control are flowed, the C3compounds producing in Calvin cycle--triose phosphate enters tenuigenin through chloroplast membranes, participate in sucrose route of synthesis, cy-FBPase catalysis fructose-1,6-diphosphate is decomposed into fructose-1, 6-diphosphate and inorganic phosphate, the generation of essential monose in regulation and control sucrose route of synthesis.
Si Lizhen etc. (03 phase of Acta Botanica Sinica.2003) are connected rice transformation by the regulating and controlling sequence of the cy-FBPase upstream region of gene of paddy rice with reporter gene, this report gene is only expressed in source tissue, this shows that fructose-1，6-diphosphatase is relevant with the metabolism of photosynthesis and sucrose in plant.The SBPase/FBPase bifunctional enzyme of blue-green algae is proceeded to potato by Masahiro Tamoi etc., it is expressed in tenuigenin, the enhancing of observing enzymic activity can promote the growth of potato side shoot, in the higher plant of enzymic activity, side shoot differentiation is more, dry weight increases, and also has similar phenotype in transgenic arabidopsis.Illustrate that the mistake expression in kytoplasm of cy-FBPase gene can affect the distribution of carbohydrate at plant materials, and then affect growing of plant.
Obtain at present spinach, sugarcane, the cDNA sequence of the fructose-1，6-diphosphatase of the several plants such as paddy rice.The reverse cDNA of potato cyFBPase is transferred to potato under the regulation and control of 35S promoter, when enzymic activity lower than wild-type 20% time, in the blade of source, accumulate 3-phoshoglyceric acid, triose phosphate and fructose-1,6-diphosphate; At light source and CO
2in sufficient situation, photosynthetic rate significantly declines.The expression that reduces cyFBPase in Arabidopis thaliana can cause the synthetic reduction of sucrose, the accumulation of phosphorylation intermediate product, and starch is synthetic to be increased.Show to play an important role in the distribution of cy-FBPase and photosynthate synthetic at regulation and control sucrose.
Sucrose is the principal mode that in higher plant, organism transports from source to storehouse, and synthetic, respiration or synthetic starch for the metabolism of cell, cell walls in the tissue of storehouse are stored up, and maintain the normal physiological metabolism of plant and growth and development process.Therefore the synthesis rate of sucrose directly has influence on the growth of plant.
The raising of soluble sugar content in vegetable cell, can improve the osmotic pressure of cell to a certain extent, maintains the stability of cell under drought stress and then strengthens drought-enduring, the Salt And Alkali Tolerance of plant, cryophylactic ability.The synthetic of sucrose not only can be improved growing of plant, also can alleviate to a certain extent the injuries of severe environment to plant such as arid, saline and alkaline, low temperature.Therefore, improving artificially the content of soluble sugar in vegetable cell, may be the effective way that improves the resistance such as drought tolerance in plants, Salt And Alkali Tolerance, anti-low temperature.
Summary of the invention
The object of the invention is to provide a kind of albumen relevant to drought resistance in plants.
Another object of the present invention is to provide the encoding gene of the above-mentioned albumen relevant to drought resistance in plants.
The present invention's the 3rd object is to provide the expression vector that contains above-mentioned encoding gene.
The present invention's the 4th object is to provide the transgenic plant that contain above-mentioned encoding gene tissue.
The present invention's the 5th object is to provide the transgenic plant that contain above-mentioned encoding gene.
The present invention's the 6th object is to provide above-mentioned encoding gene in the purposes improving on plant biomass.
The present invention's the 7th object is to provide above-mentioned encoding gene in the purposes improving on drought resistance in plants.
The present invention's the 8th object be to be provided for to increase primer of above-mentioned encoding gene.
The present invention's the 9th object is to provide the test kit that detects above-mentioned transgenic plant.
Realize technical scheme of the present invention as follows:
An albumen relevant to drought resistance in plants, be following (a) or (b) composition albumen:
(a), formed by the aminoacid sequence shown in SEQ ID No:1;
(b), produced through disappearance, replacement, the interpolation of one or several amino-acid residue by aminoacid sequence shown in SEQ ID No:1 and there is 90% above homology with SEQ ID No:1, and the aminoacid sequence with drought tolerance in plants sexual function forms.
Albumen in above-mentioned albumen (a) is rape cytoplasm fructose-1，6-diphosphatase (cy-FBPase).
The encoding gene of the above-mentioned albumen relevant to drought resistance in plants, described encoding gene is following (c) or (d):
(c) formed by the nucleotide sequence shown in SEQ ID No:2;
(d) produced through one or several Substitution, disappearance, interpolation by the nucleotide sequence shown in SEQ ID No:2, and the nucleotide sequence of coding and drought resistance in plants identical function protein composition.
Gene described in above-mentioned encoding gene (d) is made up of the nucleotide sequence shown in SEQ ID No:4.
Nucleotide sequence shown in above-mentioned SEQ ID No:4 is by rape cytoplasm fructose-1, the sudden change of three gene locuss of 6-diphosphatase gene (being abbreviated as: cy-fbpase) and come, respectively: on 173bp site, become A from G, become AAA by codon by AAG; On 258 sites, by becoming G, become AAG by codon by AAA; On the 374th site, become G by A, become GGG by corresponding codon by GGA; Although codon changes, its coded amino acid does not change.The object of these site mutations is mainly that the restriction enzyme site containing on goal gene is cut open in expression vector establishment process.
Described plant refers to corn, paddy rice, wheat, tomato, tobacco, soybean or cotton etc.
The expression vector that contains above-mentioned encoding gene.
Carrier described in above-mentioned expression vector refers to pBI121 etc.
The transgenic plant tissue that contains above-mentioned encoding gene.
The transgenic plant that contain above-mentioned encoding gene.
Above-mentioned encoding gene is in the application improving on plant biomass.
Above-mentioned encoding gene is in the application improving on drought resistance in plants.
Described plant refers to corn, paddy rice, wheat, tomato, tobacco, soybean or cotton etc.
For the primer of the above-mentioned encoding gene that increases, formed by the nucleotide sequence shown in SEQ ID No:7 and SEQ ID No:8.
For detection of the test kit that turns above-mentioned encoding gene plant, the pcr amplification primer that contains the nucleotide sequence composition shown in SEQ ID No:7 and SEQ ID No:8.
The advantage that the present invention has and beneficial effect: the present invention expresses cy-fbpase gene by mistake and improved the drought tolerance of plant.In the time that arid is processed 20 days, the sucrose content that turns cy-fbpase genetic tobacco plant all obviously raises, and root sucrose content rising amplitude maximum, can reach 10 times of left and right that contrast strain under normal condition, and plant drought tolerance strengthens; Turn sucrose content in cy-fbpase gene plant blade and improve approximately 59% than wild-type, but its starch content reduces approximately 14% than wild-type, the ratio maximum of its sucrose/starch.The enzymic activity of cy-FBPase is maximum to the sucrose content contribution of stem, and its enzymic activity is higher, and the sucrose content in stem raises more remarkable.The plant height, leaf area, stem that turn cy-fbpase gene plant are thick, dry weight/fresh weight, the every growth indexes such as number enters of blooming all have a clear superiority in than wild-type.The present invention is for utilizing genetically engineered raising plant biomass and drought tolerance that effective way is provided.
Brief description of the drawings
Fig. 1, be the pcr amplification electrophoretogram of rape cy-fbpase gene conserved sequence; Wherein 1 is Marker, and 2 is cy-fbpase gene conserved sequence.
Fig. 2, be the pcr amplification electrophoretogram of rape cy-fbpase full length gene; Wherein 1 is Marker, and 2 is rape cy-fbpase gene.
Fig. 3, be cy-fbpase gene cDNA sequence pcr amplification electrophoretogram; Be wherein 1Marker, the 2 cDNA sequences that are cy-FBPase.
Fig. 4, be the structural representation of the expression vector pGBI-fbpase of cy-fbpase gene.
Fig. 5, be that expression vector transforms the pcr amplification electrophoretogram after Agrobacterium; Wherein 1 is Marker, and 2-5 is cy-fbpase gene.
Fig. 6, be the pcr amplification electrophoretogram of transgene tobacco cy-fbpase gene; Wherein 1 and 18 is Marker, and 2-15 is Transformation of tobacco seedling, and 16 for representing positive plasmid contrast, and 17 is wild-type tobacco (negative control), and 8 and 11 do not have successful tobacco seedling for transforming.
The cy-FBPase enzymic activity column diagram of Fig. 7, transfer-gen plant and wild-type plant; Wherein 1,2 represent two different transgenic lines, and 3 represent wild-type plant.
Fig. 8, the photo of growing 10 days under artificial drought conditions for seedling for transgenosis T1; Wherein 1-6 is wild-type plant, and 7-12 is transfer-gen plant.
Fig. 9, be the graphic representation that arid is processed different time plant leaf, root sucrose content.
Figure 10, be arid plant photo before treatment; Wherein 1 is transfer-gen plant, and 2 is wild-type plant.
Figure 11, be that arid is processed the plant photo of 10 days; Wherein 1 is transfer-gen plant, and 2 is wild-type plant.
Embodiment
With specific embodiment explanation the present invention, limit the scope of the invention but be different from below.If no special instructions, related method is this area common method, and as referring to " molecular cloning " etc., or the reagent providing according to manufacturer or test kit specification sheets carry out.
The acquisition of embodiment 1, rape cy-fbpase gene fragment
Carry out as follows:
(1), the clone of cy-fbpase gene: first by known cy-fbpase DNA homolog sequence alignment, find out the conserved sequence of this gene, design primer.Conserved sequence amplified primer is as follows:
F-1 GTTGTTTTTGATCCACTTGATGG（Seq ID No:5）
R-1 GGCTTGCTCCATCAAGAACG（Seq ID No:6）
Taking rape genomic dna as template, taking F-1 and R-1 as primer, (primer is synthetic by Shanghai Sheng Gong biotechnology company limited) carries out pcr amplification, reaction system is as follows: 10 × Taq DNA polymerase buffer, 5 μ L, dNTP Mix(10mM) 2 μ L, F-1 (10 μ M) 1 μ L, R-1 (10 μ M) 1 μ L, rape genomic dna (50ng/ μ L) 1 μ L, Taq DNA polysaccharase (Taq archaeal dna polymerase is purchased from TaKaRa company, lower same) 0.5 μ L, ddH
2o 39.5 μ L, totally 50 μ L.PCR reaction conditions: 94 DEG C of 5min; 94 DEG C of 30s, 56 DEG C of 30s, 72 DEG C of 1min, 30 cycles; 72 DEG C of 8min.PCR product electrophoresis detection: the sepharose of preparation 1%, 100V voltage electrophoresis 30-40min, gel imaging is observed electrophoresis result (Fig. 1), is shown as the DNA fragmentation of 971bp size.Object band is cut, reclaim (sepharose DNA reclaims test kit purchased from Axygen company, operates according to product description, lower same), deliver Huada Gene Research Center, Beijing and check order, its sequence length is 971bp.
(2) conserved sequence obtaining is compared in the rape est sequence storehouse in NCBI search, infers the total length primer to gene.Its full length gene primer is as follows:
FBP-F 5’-ATGGATCACGAAGCAGATGC-3’ （Seq ID No:7）
FBP-R 5’-CTATTCGTCCGCAGCATACAG-3’ （Seq ID No:8）
Taking rape genomic dna as template, carry out pcr amplification taking FBP-F and FBP-R as primer; Its reaction system (50 μ L) is: 10 × Taq DNA polymerase buffer, 5 μ L, dNTP Mix(10mM) 2 μ L, FBP-F (10 μ M) 1 μ L, FBP-R (10 μ M) 1 μ L, rape genomic dna (50ng/ μ L) 1 μ L, Taq archaeal dna polymerase 0.5 μ L, ddH
2o 39.5 μ L.PCR reaction conditions: 94 DEG C of 5min; 94 DEG C of 30s, 60 DEG C of 30s, 72 DEG C of 1min20s, 30 circulations; 72 DEG C of 8min.Electrophoresis detection: the sepharose of preparation 1%, 100V voltage electrophoresis 30-40min, gel imaging is observed, and electrophoresis result (see figure 2) shows that obtaining size is the DNA fragmentation of 1910bp, and object band is cut, and reclaims.Deliver Huada Gene Research Center, Beijing and check order, the cy-fbpase gene order total length 1910bp (seeing SEQID No:3) of gained.
(3) reverse transcription
Extract total RNA(EASYspin plant RNA rapid extraction test kit of rape purchased from white Bioisystech Co., Ltd of Yuanping City, operate according to product description), and (Rever Tra Ace-α-reverse transcription test kit is purchased from TOYOBO company to carry out reverse transcription, operate according to product description), obtain single stranded DNA, taking the single stranded DNA of reverse transcription gained as template, carry out pcr amplification taking FBP-F and FBP-R as primer, electrophoresis detection (see figure 3), the gene fragment that acquisition total length is 1017bp, obtain the cDNA of cy-FBPase, delivering Huada Gene Research Center, Beijing checks order, the cy-fbpase coding gene sequence of gained is that 1017bp(is shown in Seq ID No:2), the aminoacid sequence of its coding is 338 amino acid (seeing Seq ID No:1).
The structure of embodiment 2, cy-fbpase expression vector
Carry out as follows:
(1), be cut open in vector construction process for fear of the restriction enzyme site containing on goal gene, the cDNA sequence of rape cy-fbpase gene is analyzed by software Vector NT1, to on rape cy-fbpase gene 173bp site, become A from G, on the 258th site by becoming G, on the 374th site, become G by A, although codon changes, its coded amino acid does not change, this three point mutation is by taking rape cDNA as substrate, carry out rite-directed mutagenesis with stack extension PCR method, after rite-directed mutagenesis, institute's calling sequence is shown in that SEQ IDNo:4(still claims this gene to be below: cy-fbpase).
(2), the structure of intermediate carrier: (restriction enzyme is all purchased from Fermentas company to use Pst I and Xho I, lower same) while enzyme cuts vector plasmid pG4AB and two ends have added restriction enzyme site and have been connected to the cy-fbpase gene fragment on T carrier, reclaims respectively the carrier segments of about 4kb and the gene fragment of about 1.2kb.And gene fragment is connected with carrier, build intermediate carrier pG4A-cyfbpase.
(3), by build intermediate carrier pG4A-cyfbpase and vector plasmid pBI121 through BamH I and EcoR I double digestion, reclaim the pBI121 carrier segments of about 11k and the intermediate carrier expression cassette fragment of 1.7kb left and right, the carrier segments reclaiming is connected with destination gene expression box fragment, builds to obtain plant expression vector pGBI-cy-fbpase.The expression vector structure (see figure 4) of gained, has CaMV35S promotor and Ω sequence at 5 ' end of gene, and 3 ' end has PolyA sequence and Nos terminator.
The conversion of embodiment 3, cy-fbpase gene
Carry out as follows:
The expression vector pGBI-cy-fbpase building is transformed to Agrobacterium by electrization
(1), in-80 DEG C of refrigerators, take out the LBA4404 competent cell for preparing, in 50 μ L competent cells, add to build in 1 μ L embodiment 2, plant over-express vector pGBI-cy-fbpase mixes, ice bath 15min.
(2), suck in 2mm electric shock cup (avoiding producing bubble) the 2500V conversion of shocking by electricity.
(3), electricity has swashed immediately in shocking by electricity glass and has added LB liquid nutrient medium 800 μ l; After mixing several times with rifle head, sucking-off adds in EP pipe, until the complete sucking-off of bacterium liquid, then at 28 DEG C, 180rpm cultivation recovery 4～5h.
(4), by thalline coat containing 50 μ g/mL Rifampins (referred to as: Rif), kantlex (referred to as: on LB flat board Kan), 28 DEG C be inverted cultivate within two days, can there is bacterium colony.
(5), picking list colony inoculation is in the LB liquid nutrient medium that contains 50 μ g/mLRif, Kan, bacterium liquid has shaken laggard performing PCR qualification, qualification result (see figure 5) obtains the gene fragment of 1017bp, illustrates that cy-fbpase expression vector successfully proceeds in Agrobacterium.
The acquisition of embodiment 4, cy-fbpase transfer-gen plant
(1), the acquisition of cy-fbpase transfer-gen plant
(1), Agrobacterium is cultivated single bacterium colony that picking contains goal gene cy-fbpase from LB flat board, be inoculated into (Rif 50 μ g/ml, Kan100 μ g/ml) in LB liquid nutrient medium 3ml, on constant-temperature table 28 DEG C, 180rpm shake training spend the night to OD600 be 0.6-0.8, obtain the bacterium liquid of activation.
(2), the bacterium liquid having activated is inoculated in LB liquid nutrient medium (Rif50 μ g/ml, Kan100 μ g/ml) in the ratio of 1:1000 (500 μ l:500ml ratio), 28 DEG C, 250rpm shakes bacterium and spends the night, and to the about 0.6-0.8 of bacterium liquid OD value, both can be used for transforming.4 DEG C of bacterium liquid, 4000rpm, centrifugal 5min, discard LB liquid nutrient medium, with 2 times of volume MS liquid suspensions, for infecting.
(3), infect: on Bechtop, taking 1-2 week seedling age the seedling leaf of aseptic tobacco NC89 as experiment material, the blade that cuts most advanced and sophisticated and base portion is cut into the square of 0.5cm left and right, immerse about 20min in bacterium liquid, blot the bacterium liquid on blade with aseptic filter paper, be seeded in common substratum (moiety is: MS+2mg/L6-BA+0.5mg/LIAA) upper, and all put up and down an aseptic filter paper at blade.
(4), cultivate altogether: seal culture dish with sealed membrane, in 28 DEG C of dark, cultivate 2-4 days.
(5), select to cultivate: the explant of common cultivation is transferred to screening culture medium (moiety is: MS altogether culture medium+500mg/LCar+200mg/LKan) upper, under illumination is 2,000Lux, 25-28 DEG C, 16/8h light dark condition, select to cultivate.Leaf plate edge is gently pressed in substratum, to increase selective pressure.Switching once weekly.
(6), root culture: after about 2-3 week, when resistant buds grows to 2cm left and right, cut resistant buds and transfer on root media (moiety is: 1/2MS substratum+500mg/LCep+200mg/LKan) and carry out root culture, 1-2 grows adventive root after week.
(7), the transplanting of aseptic seedling: when root grew in approximately 2 weeks, sealed membrane is opened to half in incubator to opening gradually whole hardenings.After one week, transplant seedlings in the little basin that fills vermiculite, upper cover thin film, with moisturizing, is cultivated under room temperature (25 DEG C) natural lighting, after one week, film is opened gradually.Seedling can be moved on to self-sow in the large basin that fills Nutrition Soil and vermiculite 3:1 mixed preparing when seedling survives while growing to 10cm left and right completely, transformation seedlings is detected to (T0 generation).When ripe, gather in the crops T1 for seed.
(2), the detection of cy-fbpase transfer-gen plant
Taking the genomic dna of Transformation of tobacco seedling as template, carry out pcr amplification taking FBP-F and FBP-R as primer, described primer is:
FBP-F 5’-ATGGATCACGAAGCAGATGC-3’ （Seq ID No:7）
FBP-R 5’-CTATTCGTCCGCAGCATACAG-3’ （Seq ID No:8）
PCR reaction system (totally 50 μ L): dNTP 2 μ l, 10 × Buffer, 5 μ l, FBP-F 1 μ l, FBP-R 1 μ l, the genomic dna 1 μ L of Transformation of tobacco seedling, Taq 0.5 μ L, ddH
2o 39.5 μ L.PCR reaction conditions: 94 DEG C of 5min; 94 DEG C of 30s, 60 DEG C of 30s, 72 DEG C of 60s, 35 circulations; 72 DEG C of 5min.Electrophoresis detection, amplification (see figure 6) shows the cy-fbpase gene fragment that has obtained 1017bp, illustrates that cy-fbpase gene is successfully transformed in tobacco NC89, has successfully obtained cy-fbpase genetic tobacco transformation seedlings.
The cy-FBPase enzyme biopsy test of embodiment 5 transgene tobaccos is tested
(1) test materials: the cy-fbpase genetic tobacco transformation seedlings that a. embodiment 4 obtains
B. wild-type tobacco NC89
(2) test method:
The content that discharges inorganic phosphorus in this test detection unit time in reaction system reflects enzymic activity.To wild-type tobacco, turn cyFBPase genetic tobacco cyFBPase enzymic activity detect, method is with reference to sedoheptulose-1 of Harrison etc., 7-diphosphatase (SBPase) method for detecting enzymatic activity, by substrate sedoheptulose-1 in its method, 7-bisphosphate (SBP) changes fructose-1 into, 6-bisphosphate (FBP), other steps are constant.(1) get 0.1g blade liquid nitrogen grinding, add 1ml extracting solution.At (2) 4 DEG C, the centrifugal 5min of 14,000 × g.(3) get 20 μ l supernatant liquors, add 80 μ l reaction solutions, 25 DEG C of reaction 5min.(4) add the perchloric acid termination reaction of 50 μ l 1.0M, 4 DEG C of centrifugal 10min of 14,000 × g.(5) get supernatant liquor 50 μ l and inorganic phosphorus contrast NaH
2pO
4add respectively 850 μ l ammonium molybdate solutions, react 10min at 25 DEG C.Add 150 μ l malachite green solutions (0.035% Victoria Green WPB, 0.35% PVAC polyvinylalcohol), 30min develops the color at 25 DEG C again.0-0.5mM NaH
2pO
4as standard control (6) 620nm colorimetric, with μ molg
-1s
-1represent enzymic activity.
Taking the six leaf phases of T0 generation fall two blades as material, two transgenic lines are each gets 3 strains at random, detects the enzymic activitys of cy-FBPase.The enzymic activity of the cyFBPase detecting is also that the total enzyme of this enzyme in plant lived.
Result (see figure 7) shows that the cy-FBPase enzymic activity of the transfer-gen plant of two strains has significantly improved, and is respectively 1.53 times and 1.58 times of wild-type, illustrates that activity can be expressed normally and bring into play to the cy-fbpase gene of rape in transgenic tobacco plant.The enzyme activity level of transfer-gen plant is significantly higher than and wild-type simultaneously.
The drought tolerance qualification test of embodiment 6 cy-fbpase transfer-gen plants
Carry out as follows:
(1) experiment material:
A. the cy-fbpase transgene tobacco material that embodiment 4 obtains; B. wild-type tobacco material.
(2) test method
(a) T1 is for sprouting, the screening of seed
The T1 of the cy-fbpase transgene tobacco of above-mentioned gained is sprouted on MS flat board for seed, except wild-type, the seed of transgenic line is first containing sprouting on the MS flat board of 200mg/L kantlex, screening, treat seed germination 3 days, then the tobacco seedling of normally sprouting is transferred on the PEARLITOL 50C S flat board containing 100mM and grown 5 days, then the growth of more different tobacco seedlings and the situation of taking root.
(b) simulating drought is processed the situation of taking root of lower tobacco seedling
On the PEARLITOL 50C S of 100mM flat board, tobacco seedling was grown after 10 days under artificial drought conditions, the growing state of different genetically modified tobacco seedlings.Result (see figure 8) shows that cy-fbpase transgene tobacco seedling is longer than the root of wild-type tobacco seedling, and overall growing state is better.The root growth of transgenic plant is normal, the differentiation of master root obviously, wild-type contrast shows the symptom that root growth is obstructed, especially main root extends and is obstructed, and the upper blade of transfer-gen plant is larger, entirety upgrowth situation is better than wild-type contrast, illustrates that cy-fbpase transfer-gen plant is obviously strong than wild-type plant drought tolerance.
(c) plant arid is processed the mensuration of rear blade and root sucrose content
T1 during to eight leaf phases, carries out drought stress experiment for plant strain growth, and arid is processed and watered the day before yesterday permeablely, and arid is processed first day, the tenth day, Ahau and sampled respectively once, samples altogether 3 times.Each each transfer-gen plant is all chosen 5 strains as research object, gets respectively blade and root tissue and carry out the mensuration of sucrose content.
Sucrose-determination adopts high performance liquid chromatography.Because soluble sugar is dissolved in ethanol under hot conditions, therefore adopt ethanol high temperature extraction method.The blade and the stem that take 0.1g plant are placed on the baking oven 15min of 105 DEG C, then in the baking oven of 75 DEG C, dry and spend the night to constant weight, take out and grind.Be placed in 15mL test tube, add 6～7mL80% ethanol, in 80 DEG C of water-baths, extract 30min, take out centrifugal 3000rpm5min, collect supernatant liquor.Repeat to extract twice (each 10min) centrifugal equally, collect three supernatant liquors and be incorporated in beaker, be placed in 85 DEG C of waters bath with thermostatic control, make ethanol evaporation to 2～3mL, be transferred to 50mL volumetric flask, with distilled water constant volume, for the mensuration of sucrose.
Fall three leaves as material taking eight leaf phases, analyze sucrose, the starch content of blade, and calculate sucrose starch ratio.In result transgene tobacco blade, sucrose, starch content are respectively 1.53mg/g fresh weight, 8.45mg/g fresh weight, and sucrose starch ratio is 0.18; And sucrose, starch content are respectively 0.96mg/g fresh weight, 9.82mg/g fresh weight in wild-type contrast, sucrose starch is than 0.098.The sucrose content of rotaring gene plant blade improves approximately 59% than wild-type, its starch content only has 86% of wild-type simultaneously, sucrose starch is than 1.83 times for wild-type contrast, the sucrose content and the sucrose starch ratio that turn cy-fbpase gene and can significantly improve plant leaf are described, promote the distribution of the synthetic and organic substance of sucrose in cell.
(d) arid is processed the impact on plant sucrose content
This experiment is carried out the detection of sucrose content to arid T1 after treatment for blade and the root of plant, detected result is (in table 1, what Fig. 9) arid was processed detects blade sucrose content on the 10th day, is presented between different transfer-gen plants not too large difference.But arid is after 20 days, in the blade of cy-fbpase single-gene plant, sucrose content has raising very significantly, there are some researches show at the carbohydrate of Under Drought Stress in Plant and be tending towards to root accumulation, to improve the osmotic pressure of root tissue cell, strengthen plant to arid resistivity.So measured the variation of the sucrose content of root under drought stress simultaneously, result (in table 2, Fig. 9) is presented at 20 days back root part sucrose contents of arid sharply to be increased, the more than 10 times of sucrose concentration before turning in cy-fbpase gene plant its concentration and can reaching arid and process, in wild-type plant, this value also can reach arid 3-4 before treatment doubly.It is mainly to affect root cell osmotic pressure to resist to a certain extent drought stress because of the increase of root sucrose content to arid tolerance that this presentation of results turns cy-fbpase gene plant.
Arid is processed the growing state of the plant of (seeing Figure 11) after 10 days, and the arid injury of wild-type plant is the most serious, and plant is seriously wilted; The lower blade that turns cy-FBPase single-gene tobacco plant has slight wilting dehydration symptom.
The arid plant leaf sucrose content comparison and detection test-results after treatment of table 1 (mg/g)
The arid plant root sucrose content comparison and detection test-results after treatment of table 2 (mg/g)
Note: in table, each data are means standard deviation, n=5.
These results suggest that excessively expressing and can significantly improve plant sucrose synthesis capability of cy-fbpase gene, and improve the drought tolerance of transfer-gen plant.
Claims (8)
1. an albumen relevant to drought resistance in plants, is characterized in that described albumen is made up of the aminoacid sequence shown in SEQ ID No:1.
2. the encoding gene of the albumen relevant to drought resistance in plants claimed in claim 1, is characterized in that described encoding gene is made up of the nucleotide sequence shown in SEQ ID No:2.
3. the encoding gene of the albumen relevant to drought resistance in plants claimed in claim 1, is characterized in that being made up of the nucleotide sequence shown in SEQ ID No:4.
4. according to the encoding gene described in claim 2 or 3, it is characterized in that described plant refers to tobacco.
5. contain the expression vector of the encoding gene described in claim 2 or 3.
6. the encoding gene described in claim 2 or 3 is in the application improving on drought resistance in plants; Wherein said plant refers to tobacco.
7. for the primer of the encoding gene described in the claim 2 or 3 that increases, it is characterized in that being formed by the nucleotide sequence shown in SEQ IDNo:7 and SEQ ID No:8.
8. for detection of the test kit that turns the encoding gene plant described in claim 2 or 3, it is characterized in that containing the nucleotide sequence shown in SEQ ID No:7 and SEQ ID No:8.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101942448A (en) * | 2009-07-10 | 2011-01-12 | 中国农业科学院生物技术研究所 | Novel functions of salt and drought tolerance of cotton nodulin-like gene and application thereof |
| CN102234318A (en) * | 2010-04-27 | 2011-11-09 | 中国农业科学院作物科学研究所 | Plant stress tolerance related protein TaTPRPK1, encoding gene thereof, and application thereof |
| CN102659936A (en) * | 2012-04-12 | 2012-09-12 | 中国农业科学院生物技术研究所 | Plant-stress-tolerance related protein, its encoding gene and application |
| CN102718850A (en) * | 2011-03-31 | 2012-10-10 | 中国农业科学院作物科学研究所 | Plant stress tolerance related protein GmP1 and encoding gene and application thereof |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101942448A (en) * | 2009-07-10 | 2011-01-12 | 中国农业科学院生物技术研究所 | Novel functions of salt and drought tolerance of cotton nodulin-like gene and application thereof |
| CN102234318A (en) * | 2010-04-27 | 2011-11-09 | 中国农业科学院作物科学研究所 | Plant stress tolerance related protein TaTPRPK1, encoding gene thereof, and application thereof |
| CN102718850A (en) * | 2011-03-31 | 2012-10-10 | 中国农业科学院作物科学研究所 | Plant stress tolerance related protein GmP1 and encoding gene and application thereof |
| CN102659936A (en) * | 2012-04-12 | 2012-09-12 | 中国农业科学院生物技术研究所 | Plant-stress-tolerance related protein, its encoding gene and application |
Non-Patent Citations (4)
| Title |
|---|
| Lee,J.等."GenBank Accession NO：AAZ86538.1".《GenBank》.2005,第1-2页. |
| Lee,J.等."GenBank Accession NO：DQ165552.1".《GenBank》.2005,第1-2页. |
| Lee,J.等."GenBank Accession NO:AAZ86538.1".《GenBank》.2005,第1-2页. * |
| Lee,J.等."GenBank Accession NO:DQ165552.1".《GenBank》.2005,第1-2页. * |
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