CN101503468A - Protein related to pollen germination and/or pollen tube growth, coding gene and use thereof - Google Patents
Protein related to pollen germination and/or pollen tube growth, coding gene and use thereof Download PDFInfo
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Abstract
The invention discloses a protein related to pollen grain germination and/or pollen tube growth, the encoding genes and the application thereof. The protein is the following protein shown in (1) or (2): (1) the protein being composed of amino acid sequences shown in sequence two of a sequence table; (2) the protein comprises the amino acid in the sequence two of the sequence table substituted and/or lacked and/or added and is related to the stress tolerance of plant. The protein and the encoding gene thereof related to the pollen grain germination and/ or pollen tube growth can be used for the genetic transformation research of the plant, increases the pollen germination rate of plant and/or the growth speed of the pollen tube and the reproductive efficiency of the plant, shortens the breeding process, has important theoretical and practical actual meaning for adjusting and controlling the reproductive development and gene engineering breeding of woody plants with slow growth development and accelerating the genetic improvement of the woody plants.
Description
Technical field
The present invention relates to a kind of albumen relevant and encoding gene and application with pollen germination and/or pollen tube growth.
Background technology
Pollen germination, pollen tube growth are the amphigenetic important steps of spermatophyte.Spermatophyte must rely on pollen tube and transmit sperm and ovum fusion, finishes fertilization.Metabolic activity in the pollen tube is very active, and very high protein, lipid and carbohydrate synthesis rate are arranged, and these materials all participate in the synthetic etc. of the top expansion of pollen tube and new cell walls.Pollen tube is typical apical growth cell, and its growth has directivity, thereby pollen tube provides a kind of special modular system for experimental study.More than the result of study of cytoskeleton in pollen and the pollen tube, get for experiment material observation post with the angiosperm, gymnosperm is owing to exist difference with angiospermous evolution, pollen development and sperm production process are more complicated, relate to other several times mitotic division, therefore, the distribution of gymnosperm inner cell skeleton is also different.
Microtubule is made up of the tubulin subunit of two types of α and β, two types tubulin subunit forms the tubulin dimer, the tubulin dimer is the fundamental unit of microtubule assembling, the elongated tubular organoid structure that microtubule is made up of the tubulin dimer.Tubulin has important effect in the sprouting of plant pollen and polar growth, the research for the effect of microtubule in pollen germination and pollen tube growth at present only limits to protein level, and does not study from the gene transcription regulation level.
Summary of the invention
The purpose of this invention is to provide a kind of albumen and the encoding gene thereof relevant with pollen germination and/or pollen tube growth.
The albumen relevant with pollen germination and/or pollen tube growth provided by the present invention, name is called PwTUA1, derive from the blue or green bar (Picea wilsorii Mast-Pmastersii Mayr.) of Pinaceae, Picea, is following 1) or 2) protein:
1) protein of forming by the aminoacid sequence shown in the sequence in the sequence table 2;
2) with the aminoacid sequence of sequence in the sequence table 2 through the replacement of one or several amino-acid residue and/or disappearance and/or interpolation and relevant with pollen germination and/or pollen tube growth by 1) deutero-protein.
The sequence of table 1. label
| Label | Residue | Sequence |
| Poly-Arg | 5-6 (being generally 5) | RRRRR |
| Poly-His | 2-10 (being generally 6) | HHHHHH |
| FLAG | 8 | DYKDDDDK |
| Strep-tag?II | 8 | WSHPQFEK |
| c- |
10 | EQKLISEEDL |
Above-mentioned 2) but in the PwTUA1 synthetic, also can synthesize its encoding gene earlier, carry out biology again and express and to obtain.Above-mentioned 2) encoding gene of the PwTUA1 in can be by the codon that lacks one or several amino-acid residue in the dna sequence dna shown in the 5 ' terminal 4-1356 bit base with sequence in the sequence table 1, and/or carry out the missense mutation of one or several base pair, and/or obtain at the encoding sequence that its 5 ' end and/or 3 ' end connects the label shown in the table 1.
Above-mentioned cDNA gene with pollen germination and/or pollen tube growth associated protein also belongs to protection scope of the present invention.
Specifically can be following 1 with the cDNA gene of pollen germination and/or pollen tube growth associated protein)-4) in arbitrary described gene:
1) its encoding sequence be in the sequence table sequence 1 from 5 ' terminal 4-1356 position deoxyribonucleotide;
2) its nucleotide sequence is the sequence 1 in the sequence table;
3) under stringent condition, can hybridize with the dna sequence dna that sequence in the sequence table 1 limits and the dna molecular above-mentioned and pollen germination and/or pollen tube growth associated protein of encoding;
4) with 1) gene have the homology more than 90% and the above-mentioned proteic dna molecular relevant of encoding with pollen germination and/or pollen tube growth.
Gene in the described step 4) is with 1) gene homology more than 95% is preferably arranged.
Above-mentioned stringent condition can be at 6 * SSC, in the solution of 0.5%SDS, 65 ℃ of hybridization down, uses 2 * SSC then, and 0.1%SDS and 1 * SSC, 0.1%SDS respectively wash film once.
Increase above-mentioned PwTUA1 full length gene or arbitrary segmental primer to also belonging to protection scope of the present invention.
Contain above-mentioned recombinant expression vector, expression cassette, transgenic cell line and reorganization bacterium and also belong to protection scope of the present invention with pollen germination and/or pollen tube growth associated protein encoding gene.
Available existing plant expression vector construction contains the recombinant expression vector of PwTUA1 gene.Described plant expression vector comprises the double base agrobacterium vector and can be used for the carrier etc. of plant micropellet bombardment, as pCAMBIA3301, pCAMBIA1300, pBI121, pBin19, pCAMBIA2301, pCAMBIA1301-UbiN or other plant expression vector of deriving.The plant expression vector that carries the present invention and pollen germination and/or pollen tube growth associated protein encoding gene PwTUA1 can be led by Ti-plasmids, Ri plasmid, plant viral vector, directly DNA conversion, microinjection, electricity, particle bombardment or conventional biological method such as agriculture bacillus mediated are transformed in vegetable cell or the tissue.By the plant transformed host both can be angiosperms such as Arabidopis thaliana, also can be gymnosperms such as blue or green bar.
When using the gene constructed recombinant plant expression vector of PwTUA1, before its transcription initiation Nucleotide, can add any enhancement type, composing type, organizing specific type or inducible promoter, as cauliflower mosaic virus (CAMV) 35S promoter, general living plain gene Ubiquitin promotor (pUbi) etc., they can use separately or be used in combination with other plant promoter; In addition, when using gene constructed plant expression vector of the present invention, also can use enhanser, comprise translational enhancer or transcriptional enhancer, these enhanser zones can be ATG initiator codon or neighboring region initiator codon etc., but must be identical with the reading frame of encoding sequence, to guarantee the correct translation of whole sequence.The source of described translation control signal and initiator codon is widely, can be natural, also can be synthetic.Translation initiation region can be from transcription initiation zone or structure gene.
For the ease of transgenic plant cells or plant being identified and screening, can process used plant expression vector, can in plant, express enzyme or the gene (gus gene, luciferase genes etc.) of luminophor, antibiotic marker thing (gentamicin marker, kantlex marker etc.) or the anti-chemical reagent marker gene (as anti-weedkiller gene) etc. that can produce colour-change with resistance as adding.
Another object of the present invention provides a kind of method of cultivating the transgenic plant of pollen germination and/or the raising of pollen tube growth speed.
The method of the transgenic plant that cultivation pollen germination provided by the present invention and/or pollen tube growth speed improve, be that above-mentioned encoding gene PwTUA1 with pollen germination and/or pollen tube growth associated protein is imported in the plant, obtain the transgenic plant that pollen germination and/or pollen tube growth speed improve.
Described plant can be angiosperms such as Arabidopis thaliana, also can be gymnosperms such as blue or green bar.
The present invention clones the PwTUA1 gene and is connected on the expression vector from blue or green bar, utilize particle bombardment and Agrobacterium infestation method that this gene is imported in blue or green bar pollen and the Arabidopis thaliana pollen respectively.Experimental result shows that the germination rate of the blue or green bar pollen of the transgenosis that is obtained can reach 92.4 ± 12%, and the germination rate of the transgenic arabidopsis pollen that is obtained can reach 88 ± 6%; The speed of growth of the blue or green bar pollen tube of transgenosis can reach 13.7 ± 1.38 μ m/h, and the speed of growth of transgenic arabidopsis pollen tube can reach 63.3 ± 8.7 μ m/h.Coerce under the environment at boron ion and calcium ion, transgenic arabidopsis pollen tube growth speed shows advantage than wild-type or the Arabidopis thaliana pollen tube that changes empty carrier over to.When containing 0.1g/L boric acid in the substratum, the transgenic arabidopsis pollen tube growth speed that changes empty carrier over to is 20.0 ± 2.9 μ m/h, and the transgenic arabidopsis pollen tube growth speed that changes recombinant expression vector PBI121-1-PwTUA1 over to is 42.7 ± 3.7 μ m/h; When containing the 20mM calcium ion in the substratum, the transgenic arabidopsis pollen tube growth speed that changes empty carrier over to is 16.7 ± 2.3 μ m/h, and the transgenic arabidopsis pollen tube growth speed that changes recombinant expression vector PBI121-1-PwTUA1 over to is 35.6 ± 3.1 μ m/h.Albumen of the present invention is compared in GenBank, and the result shows, the homology of the tubulin in the genome of this albumen and the Arabidopis thaliana of having announced, paddy rice, cotton and willow is all than higher, and this albumen has the characteristic of single-minded expression in the pollen.The Study on Genetic Transformation that albumen relevant with pollen germination and/or pollen tube growth of the present invention and encoding gene thereof can be used for plant, improve the pollen germination rate of plant and/or pollen tube growth speed, early flowering, raising plant reproductive efficiency, shorten breeding process, the genetic improvement that carries out reproductive development regulation and control, genetic engineering breeding, acceleration woody plant for the woody plant slowly of growing has important theory and practical significance.
Description of drawings
Fig. 1 is the expression of PwTUA1 gene in each tissue of blue or green bar
Fig. 2 is the measurement result that changes empty carrier over to and change the blue or green bar pollen germination rate of PwTUA1 gene over to
Fig. 3 is the measurement result that changes empty carrier over to and change the blue or green bar pollen tube growth speed of PwTUA1 gene over to
Fig. 4 is the measurement result that changes empty carrier over to and change the Arabidopis thaliana pollen germination rate of PwTUA1 gene over to
Fig. 5 is the measurement result that changes empty carrier over to and change the Arabidopis thaliana pollen tube growth length of PwTUA1 gene over to
Embodiment
Experimental technique described in the following embodiment if no special instructions, is ordinary method; Described reagent and biomaterial if no special instructions, all can obtain from commercial channels.
1, the acquisition of the intermediate sequence of gene PwTUA1
In blue or green bar (Zhang LY, Lin JX, et al.Plant Science, 2007,172:1210-1217.) inflorescence open fine morning, the tweezers with sterilizing take off the xanchromatic pollinium, be placed on the clean paper of white, put into the dry glass bottle of sterilizing then, place indoor drying in the shade, be sub-packed in again in the exsiccant vial, sealing places in-20 ℃ of refrigerators storage standby.Extract total RNA of blue or green bar pollen, reverse transcription obtains cDNA, with this cDNA is template, high conservative region design degenerated primer 5 '-GATGCHTTCWACACHTTYTTYAG-3 ' and 5 '-AGGGCRGCVAGRTCCTCACG-3 ' according to the tubulin encoding gene of the Arabidopis thaliana of having announced among the GenBank, paddy rice, cotton and willow carries out pcr amplification.In the designed primer, H represents A, T or C, and W represents A or T, and Y represents C or T, and R represents A or G, and V represents G, A or C.
PCR reaction conditions: 94 ℃ of pre-sex change 5min of elder generation; 94 ℃ of 30sec then, 56 ℃ of 30sec, 72 ℃ of 1min, totally 35 circulations; 72 ℃ are extended 5min again.
Reclaim above-mentioned PCR reaction product, carry out 1.0% agarose gel electrophoresis and detect, the result obtains the band of about 879bp, i.e. the intermediate sequence of gene PwTUA1.
2, the acquisition of 3 ' RACE
Intermediate sequence design 3 ' RACE primer: 5 '-CTGGAACACACCGATGTGGC-3 ' of the gene PwTUA1 that obtains according to above-mentioned steps 1 utilizes 3 ' PCR primer: 5 '-GTATCGATGCCCACCCTCTAGAGGCCGAGGCGGCCGACA-3 ' and above-mentioned 3 ' RACE primer to carry out pcr amplification.
PCR reaction conditions: 94 ℃ of pre-sex change 5min of elder generation; 94 ℃ of 30sec then, 62 ℃ of 30sec, 72 ℃ of 1min, totally 35 circulations; 72 ℃ are extended 5min again.
Reclaim above-mentioned PCR reaction product, carry out 1.0% agarose gel electrophoresis and detect, the result obtains the band of about 774bp, i.e. 3 ' the terminal sequence of gene PwTUA1.
3, the acquisition of 5 ' RACE
Intermediate sequence design 5 ' RACE primer: 5 '-GAACCGAGACCAGAACCAGTTCC-3 ' of the gene PwTUA1 that obtains according to above-mentioned steps 1,5 ' PCR primer: 5 '-TTCCACCCAAGCAGTGGTATCAACGCAGAGTGG-3 ' and above-mentioned 5 ' RACE primer carry out pcr amplification.
PCR reaction conditions: 94 ℃ of pre-sex change 5min of elder generation; 94 ℃ of 30sec then, 62 ℃ of 30sec, 72 ℃ of 1min, totally 35 circulations; 72 ℃ are extended 5min again.
Reclaim above-mentioned PCR reaction product, carry out 1.0% agarose gel electrophoresis and detect, the result obtains the band of about 452bp, i.e. 5 ' the terminal sequence of gene PwTUA1.
4, the acquisition of total length PwTUA1 gene cDNA sequence
For obtaining the cDNA sequence of total length PwTUA1 gene, respectively 5 ' RACE that amplifies in step 3 and step 2 and 3 ' RACE sequence two ends design primer 5 '-ATGAGAGAGTGCATCTCGATCCAC-3 ' and 5 '-TCAGTACTCGCCTCCCTCATCACC-3 ', be that template is carried out pcr amplification with blue or green bar pollen cDNA.
PCR reaction conditions: 94 ℃ of pre-sex change 5min of elder generation; 94 ℃ of 30sec then, 56 ℃ of 30sec, 72 ℃ of 90sec, totally 35 circulations; 72 ℃ are extended 5min again.
Reclaim above-mentioned PCR reaction product, carry out 1.0% agarose gel electrophoresis and detect, the result obtains the band of 1356bp, reclaims this band and is connected in the pMD18-T cloning vector and carries out sequencing.Sequencing result shows that the nucleotide sequence of the total length PwTUA1 gene of acquisition is shown in sequence in the sequence table 1, and its amino acid sequence coded is shown in sequence in the sequence table 2.Utilize BLAST software that the encoding gene of the tubulin of Arabidopis thaliana, paddy rice, cotton and the willow of having announced among the total length PwTUA1 gene of above-mentioned acquisition and the GenBank is carried out homology relatively, the result shows that its homology is for being respectively 88.47%, 96.9%, 97.78% and 99.78%.。
Extract the RNA of blue or green bar pollen, needle, bark and root tissue respectively, cDNA is synthesized in reverse transcription; Being template with above-mentioned each cDNA that organizes respectively then, is that primer carries out pcr amplification with 5 '-TGCATGATCTCCAATTCGAC-3 ' and 5 '-GCACTGTTCTCAACATGAAG-3 ', detects the expression of PwTUA1 gene in blue or green bar different tissues.While, as confidential reference items, 5 ' primer of amplification EF1-α gene is: 5 '-AACTGGAGAAGGAACCCAAG-3 ', 3 ' primer was: 5 '-AACGACCCAATGGAGGATAC-3 ' with EF1-α gene.
PCR reaction conditions: 94 ℃ of pre-sex change 5min of elder generation; 94 ℃ of 10sec then, 56 ℃ of 10sec, 72 ℃ of 20sec, totally 28 circulations; 72 ℃ are extended 5min again.
The expression of PwTUA1 gene in each tissue as shown in Figure 1.The result shows that having only has the PwTUA1 expression of gene in the pollen, and this gene not in its hetero-organization.
Embodiment 3, cultivation pollen germination rate and blue or green bar pollen of the fast transgenosis of pollen tube growth and Arabidopis thaliana
1, the structure of expression vector
With plasmid PBI121 (available from Clontech company) with contain the plasmid pLat52-7 (DavidTwell of Lat52 promotor, Rod Wing, J udy Yamaguchi et al.Isolation and expresstion of an anther-specificgene from tomato, Mol Gen Genet, 1989,217:240～245; David Twell, Theodore MKlein, Michael E Fromm et al.Transient expression of chimeric genes delivered intopollen by microprojectile bombardment.Plant Physiol, 1989,91:1270～1274.) carry out double digestion with restriction enzyme HindIII and BamHI respectively, then enzyme is cut product and be connected and obtain recombinant vectors PBI121-1; The PwTUA1 gene of getting 1 acquisition of 2 μ l the foregoing descriptions is connected with the pMD18-T carrier, and operation steps is carried out according to the specification sheets of the product pMD18-T Vector system of Promega company; The connection product that obtains is carried out double digestion with restriction enzyme XbaI and BamHI, and enzyme is cut product and is connected with the plasmid PBI121-1 that cuts through same enzyme; Connect product transformed into escherichia coli DH5 α competent cell, and be coated on overnight incubation on the LB flat board that contains 5-bromo-4-chloro-3-indoles-β-D-galactoside, X-gal and 100ug/ml penbritin.The single bacterium colony of picking white, overnight incubation and carry out bacterium colony PCR and identify in the LB liquid nutrient medium; Alkaline process extraction plasmid DNA is carried out sequencing simultaneously.Sequencing result shows that the PwTUA1 gene correctly is connected on the plasmid PBI121-1, with the recombinant expression vector called after PBI121-1-PwTUA1 that obtains.
2, particle bombardment instantaneous conversion PwTUA1 gene is in blue or green bar pollen and the mensuration of transgenosis pollen germination rate and pollen tube growth speed
Blue or green bar pollen (is contained 15% quality percentage composition sucrose, 0.03% quality percentage composition nitrocalcite, 0.01% quality percentage composition boric acid and 5mM citric acid-phosphoric acid buffer with substratum, pH 5.8) suspension culture is in culture dish middle part, forms the round spot of diameter 2cm.Respectively the recombinant expression vector PBI121-1-PwTUA1 that the above-mentioned steps 1 of 1ug, 2.5ug and 5ug is made up with the tungsten powder parcel after, utilize the blue or green bar pollen of the above-mentioned cultivation of PDS-1000/He particle gun (available from U.S. Bio-Rad company) bombardment.Simultaneously in contrast with blue or green bar pollen that does not change any plasmid over to and the blue or green bar pollen that changes plasmid PBI121-1 over to.
The blue or green bar pollen that the said gene rifle bombarded is sprouted down in room temperature (25 ± 1 ℃).Fluorescent microscope is observed the sprouting and the growing state of the blue or green bar pollen of transgenosis down, and when changing plasmid 12h and 24h over to, the measurement result of the germination rate of transgenosis pollen and pollen tube growth speed as shown in Figures 2 and 3.Wherein CK represents not change over to the blue or green bar pollen of any plasmid, and 0 expression changes the blue or green bar pollen of plasmid PBI121-1 over to.Three repetitions are established in experiment.
The result shows that the germination rate that changes the transgenosis pollen of recombinant expression vector PBI121-1-PwTUA1 over to improves, and germination rate can reach 92.4 ± 12%; And the pollen tube growth speed of transgenosis pollen is also obviously accelerated, and the speed of growth of transgenosis pollen tube can reach 13.7 ± 1.38 μ m/h.
3, the cultivation of transgenic arabidopsis
Transform the competent cell of Agrobacterium GV3101 with the recombinant expression vector PBI121-1-PwTUA1 of above-mentioned steps 1 structure, the mono-clonal that picking changes the Agrobacterium of recombinant expression vector PBI121-1-PwTUA1 over to is inoculated in the LB liquid nutrient medium that contains the 50mg/L kantlex, 28 ℃ of shaking culture two days.With centrifugal 5 minutes of fermented liquid 3000rpm/min, gained Agrobacterium precipitation suspended with the liquid that infects that contains 5% sucrose and 0.03%SilwetL-77.
Employing is stained with colored dip method and is transformed the environmental wild-type Arabidopis thaliana (Col-0) of Colombia, gathers in the crops this seed (T that present age, the transgenic arabidopsis plant was connect
1Generation), the seed of sprouting in the MS substratum screening that contains 50mg/L GM.The T that will on above-mentioned substratum, sprout
1Move on in the compost results seed (T for seedling
2Generation), the T that obtains isozygotying through identical screening process then
3For the transgenic arabidopsis seed.At last with T
3Directly be seeded in the compost T that grows for the transgenic arabidopsis seed
3Blooming about two weeks of growth under the long day condition for the transgenic arabidopsis plant.Simultaneously with wild-type Arabidopis thaliana that does not change any plasmid over to and the T that changes plasmid PBI121-1 over to
3For transgenic arabidopsis in contrast.
Get the above-mentioned T that changes plasmid PBI121-1 over to
3For transgenic arabidopsis and above-mentioned T
3The flower pesticide that is about to open flower that generation changes the transgenic arabidopsis of recombinant expression vector PBI121-1-PwTUA1 over to carries out the sprouting and the cultivation of pollen.Flower pesticide was at room temperature placed 2 hours, made its dehydration; Then flower pesticide is placed (consisting of of pollen germination substratum: 18g/L sucrose, 0.01g/L boric acid, 1mM sal epsom, 1mM calcium chloride, 1mM nitrocalcite and 0.5g/L agar) on the pollen germination substratum.After 6 hours,, measure the length of pollen tube by the sprouting situation of microscopic examination pollen and the growing state of pollen tube.The measurement result of the germination rate of transgenosis pollen and pollen tube growth speed as shown in Figure 4 and Figure 5.Among Fig. 4, WT represents to change over to the Arabidopis thaliana of plasmid PBI121-1, and Lat52:PwTUA1 represents to change over to the transgenic arabidopsis of recombinant expression vector PBI121-1-PwTUA1.Among Fig. 5,2 expressions change the Arabidopis thaliana of plasmid PBI121-1 over to, and 1 expression changes the transgenic arabidopsis of recombinant expression vector PBI121-1-PwTUA1 over to.Three repetitions are established in experiment.
The result shows that the germination rate that changes the transgenic arabidopsis pollen of recombinant expression vector PBI121-1-PwTUA1 over to can reach 88 ± 6%, and the speed of growth of pollen tube can reach 63.3 ± 8.7 μ m/h.When the concentration of substratum mesoboric acid is 0.1g/L, is the boron ion and coerces; When containing 10mM calcium chloride and 10mM nitrocalcite in the substratum, be calcium ion and coerce.When the boron ion is coerced, change the T of plasmid PBI121-1 over to
3For transgenic arabidopsis pollen tube growth speed is 20.0 ± 2.9 μ m/h, and the transgenic arabidopsis pollen tube growth speed that changes recombinant expression vector PBI121-1-PwTUA1 over to is 42.7 ± 3.7 μ m/h; When calcium ion is coerced, change the T of plasmid PBI121-1 over to
3For transgenic arabidopsis pollen tube growth speed is 16.7 ± 2.3 μ m/h, and the transgenic arabidopsis pollen tube growth speed that changes recombinant expression vector PBI121-1-PwTUA1 over to is 35.6 ± 3.1 μ m/h.
Sequence table
＜110〉Beijing Forestry University
Shandong Agricultural University
＜120〉a kind of albumen relevant and encoding gene and application with pollen germination and/or pollen tube growth
<130>CGGNARZ92147
<160>2
<210>1
<211>1356
<212>DNA
＜213〉blue or green bar (Picea wilsorii Mast-Pmastersii Mayr.)
<400>1
<210>2
<211>451
<212>PRT
＜213〉blue or green bar (Picea wilsorii Mast-Pmastersii Mayr.)
<400>1
Claims (10)
1, a kind of albumen is following 1) or 2) protein:
1) protein of forming by the aminoacid sequence shown in the sequence in the sequence table 2;
2) with the amino acid residue sequence of sequence in the sequence table 2 through the replacement of one or several amino-acid residue and/or disappearance and/or interpolation and relevant with pollen germination and/or pollen tube growth by 1) deutero-protein.
2, the described proteic encoding gene of claim 1.
3, encoding gene according to claim 2 is characterized in that: described proteic cDNA gene is following 1)-4) in arbitrary described gene:
1) its encoding sequence be in the sequence table sequence 1 from 5 ' terminal 4-1356 position deoxyribonucleotide;
2) its nucleotide sequence is the sequence 1 in the sequence table;
3) under stringent condition, can hybridize and the described proteic dna molecular of coding claim 1 with the dna sequence dna that sequence in the sequence table 1 limits;
4) with 1) gene have the homology 90% or more and the described proteic dna molecular of claim 1 of encoding.
4, the recombinant expression vector that contains claim 2 or 3 described genes.
5, contain claim 2 or 3 described expression of gene boxes.
6, the transgenic cell line that contains claim 2 or 3 described genes.
7, the reorganization bacterium that contains claim 2 or 3 described genes.
8, total length or arbitrary segmental primer of amplification claim 2 or 3 described genes are right.
9, a kind of method of cultivating the transgenic plant of pollen germination and/or the raising of pollen tube growth speed is that claim 2 or 3 described encoding genes are changed in the plant, obtains the transgenic plant that pollen germination and/or pollen tube growth speed improve.
10, method according to claim 9 is characterized in that: described plant is Arabidopis thaliana or blue or green bar.
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| CN108822195A (en) * | 2018-06-14 | 2018-11-16 | 南京农业大学 | Dangshan pear has albumen, encoding gene PbrTTS1 and its application for promoting pollen tube growth function |
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| CN116473082A (en) * | 2023-06-21 | 2023-07-25 | 中国农业科学院蜜蜂研究所 | Auxiliary agent for accelerating elongation of pollen tube and formula thereof |
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| CN102168060A (en) * | 2011-01-04 | 2011-08-31 | 沈阳师范大学 | Method for observing dynamic growth process of pollen tube germination and growth |
| CN108822195A (en) * | 2018-06-14 | 2018-11-16 | 南京农业大学 | Dangshan pear has albumen, encoding gene PbrTTS1 and its application for promoting pollen tube growth function |
| CN108822195B (en) * | 2018-06-14 | 2020-10-23 | 南京农业大学 | Protein with function of promoting growth of pollen tube of Dangshan pear, coding gene PbrTTS1 and application of coding gene PbrTTS1 |
| WO2022042619A1 (en) * | 2020-08-27 | 2022-03-03 | 云南大学 | Plant pollen tube growth gene and application |
| CN114149983A (en) * | 2021-12-07 | 2022-03-08 | 中国林业科学研究院 | Thioredoxin for regulating and controlling identification of plant pollen tube and stigma and preparation method thereof |
| CN116473082A (en) * | 2023-06-21 | 2023-07-25 | 中国农业科学院蜜蜂研究所 | Auxiliary agent for accelerating elongation of pollen tube and formula thereof |
| CN117362404A (en) * | 2023-09-06 | 2024-01-09 | 广东省农业科学院果树研究所 | Longan pollen with nano magnetic bead mediated overexpression of DlNIP1 transport protein and application thereof |
| CN117362404B (en) * | 2023-09-06 | 2024-06-11 | 广东省农业科学院果树研究所 | Longan pollen with nano magnetic bead mediated overexpression DlNIP transport protein and application thereof |
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