CN1934259A - Plants having improved growth characteristics and method for making the same - Google Patents

Abstract

The present invention concerns a method for improving the growth characteristics of plants by introducing and expressing in a plant a nucleic acid encoding an S3a protein. The invention also relates to transgenic plants having introduced therein a nucleic acid encoding an S3a protein, which plants have improved growth characteristics relative to corresponding wild type plants. The present invention also concerns constructs useful in the methods of the invention.

Description

Have the plant of growth characteristics of improvement and the method for preparing described plant

Relate generally to biology field of the present invention, and relate to a kind of method that improves plant growth characteristics.More particularly, the present invention relates to improve the especially method of productive rate of plant growth characteristics by the nucleic acid of introducing in plant and expression coding small subunit ribosomal protein (S3a).The invention still further relates to the plant of wherein having introduced the S3a coding nucleic acid, this plant has the growth characteristics of improvement with respect to the corresponding wild-type plant.

The supply of the ever-increasing world population and the continuous minimizing in the arable land that can be used for agricultural evokes farming research to the efficient development that improves agricultural.The traditional means of farm crop and gardening improvement utilizes breeding technique to identify to have the plant of desired characteristic.Yet such breeding technique has some shortcomings, and promptly these technology generally are labor-intensive, and produces the plant that contains heterogeneous hereditary component usually, and this is not always to produce the anticipant character that passes over from mother plant.Development of molecular biology has allowed the human germplasm of modifying animal and plant.The genetic engineering of plant need separate and operate genetic material (generally being the form of DNA or RNA), needs subsequently this genetic material is imported in the plant.This technology can provide the crop or the plant of economics, agronomy or gardening proterties with multiple improvement.A kind of proterties with special economic interests is a productive rate.Productive rate is normally defined the economic worth weighed of making deposits yields.This can be from quantity and/or the definition of quality aspect.Productive rate directly depends on Several Factors, for example, and the quantity of organ and size, plant structure (for example, branched quantity), seed production or the like.The growth of root, dietetic alimentation and stress tolerance also are the important factors of determining productive rate.Can improve crop yield by optimizing one of above-mentioned factor.

The seventies in 20th century is early stage, and the profile of rrna assembling approach has been appeared in the saccharose gradient analysis in one's mind.3 kinds of main preribosomal particles have been identified in these researchs.Early stage 90S particle (' S ' is sedimentation rate) is processed to the preceding rrna of 66S and 43S subsequently, and they are respectively the precursor of sophisticated 60S and 40S subunit.The 60S subunit is made up of the ribosome-RNA(rRNA) of 25S, 5.8S and 5S after processing.In ripening process, numerous protein will interact and produce final structure.The 40S subunit is made up of the 18S ribosome-RNA(rRNA), and before ripe mixture forms, has numerous protein to interact equally.These two kinds of subunits are assembled into big mixture, and wherein the 40S small subunit is in conjunction with mRNA and tRNAs, and big subunit catalysis peptide bond forms.About 2/3 of rrna quality is made up of RNA, and 1/3 is made up of protein.Protein is named according to the ribosomal subunit under it: small subunit (S1 to S31) and big subunit (L1 to L44).Most protein interact with common a plurality of RNA elements from the different structure territory, to constitute and stable rRNA tertiary structure.Although the crucial activity of decoding and peptide displacement is based on the process of RNA, protein plays a part positive in the function aspects that may evolve to the protein building-up process streamline.Except they typical rrna effects, many ribosomal proteins also have the non-ribosomal effect, repair as DNA.

S3a (cyc07) cloned first in 1991.It the coding 40S ribosomal protein (small subunit), with the v-fos conversion effet factor (by the fte-1 genes encoding of people and rat), TNF-α inductive mouse TU-11 dna homolog, and be similar to yeast PLC1 and PLC2.Mammals fte-1, plant cyc-07 and yeast PLC2 have shown it is cell cycle regulating, and participate in proteinic synthetic on the rrna level.

The ability of the various growth characteristics of improvement plant, to in such as production, arboriculture, Horticulture, the forestry of crop enhancing, plant breeding, ornamental plant, the field such as production (be used for the biotechnology production of material such as medicine, antibody or vaccine, or the bio-transformation of organic waste) that is used for the algae of bio-reactor and other such field, have many application.

Have now found that, in plant, introduce and express the nucleic acid of coding small subunit ribosomal protein (S3a), produced the plant that for the corresponding wild-type plant, has the growth characteristics of improvement.Therefore,, provide a kind of method that is used to improve the growth characteristics of plant, be included in the nucleic acid of introducing and expressing coding S3a in the plant according to one embodiment of the invention.

Best, by implementing method of the present invention, produce plant, the especially productive rate of Zeng Jiaing, particularly seed productive rate of growth characteristics with multiple improvement.

Be used to refer to respectively with respect to the corresponding wild-type plant as defined term in the literary composition " productive rate of increase ", in following increase aspect one or more: (i) biomass (weight) that increases of the one or more parts of plant, (can gather in the crops) part on the ground particularly, any other biomass of the part gathered in the crops of root biomass that increases or increase, the (ii) seed productive rate of Zeng Jiaing, it may come from the seed biomass (seed weight) of increase, may be the increase of every strain plant seed weight or the increase on indivedual seed basis, and the increase of described seed weight may be because of due to the seed size that changes, as seed length and/or seed width and/or seed area; (iii) (full) seed amount of Zeng Jiaing; The (iv) seed size of Zeng Jiaing, this may also influence the composition of seed; (the v) seed volume of Zeng Jiaing, this also may influence the composition of seed; (the vi) harvest index of Zeng Jiaing, it can be expressed as can gather in the crops part accounts for total biomass as the productive rate of seed ratio; (thousand seeds that vii) increase heavy (TKW) are to infer from the quantity and their gross weight of the full seed of counting.The TKW that increases may come from the seed size and/or the seed weight of increase.

According to a preferred embodiment of the invention, the increase of productive rate is contained as mentioned (ii) to the (gain in yield vii) on any one or the multinomial defined seed level.

With cereal is example, and gain in yield can show as following one or more aspect: the increase of per hectare or every acre of plant number, and the increase of every strain plant fringe, line number, every row are planted the benevolence number, grain is heavy, thousand seeds heavy, the increase of fringe length/diameter, or the like.With the rice is example, and gain in yield can show as the increase of following one or more aspects: per hectare or every acre of plant number, the No. of inflorescences of every strain plant, spikelet number on each inflorescence, spend number on each inflorescence, the increase of the full rate of seed, the increase that thousand seeds are heavy, or the like.The increase of productive rate may also cause the structure that changes, or may take place as the result of the structure that changes.

According to preferred characteristics of the present invention, implement method of the present invention and produced the plant of productive rate with increase, it shows as following at least one aspect with respect to control plant respectively: the TKW of increase, (full) seed number of increase, the harvest index of increase seed weight and increase.Therefore, according to the present invention, provide a kind of method that is used to increase plant yield, this method is included in the nucleic acid of introducing in the plant and expressing coding S3a polypeptide.

Because transgenic plant according to the present invention have the productive rate of increase, for the growth velocity of the corresponding wild-type plant of the respective stage that is in its life cycle, these plants demonstrate the growth velocity (at them during the life cycle of small part) of increase probably.The growth velocity that increases may be that one or more parts (comprising seed) of plant are specific, maybe may be whole basically plant.Have increase growth velocity plant in addition may demonstrate early flowering.The increase of growth velocity may occur in plant one or more stages of life cycle, or basically during whole plants life cycle.The plant early stage growth velocity that increases of life cycle may reflect the enhanced growing ability.The increase of growth velocity may change the harvest cycle of plant, makes that plant can be than possible situation more late sowing kind and/or early harvest more.If growth velocity fully increases, may allow to sow more kinds of sons (for example sow and gather in the crops rice plants, then sow and gather in the crops more rice plants, all these is in a traditional vegetative period) of identical plant species.Similarly, if growth velocity fully increases, may allow to sow different plant species more kinds of sons (for example sow and gather in the crops rice plants, then, for example, sowing and randomly gather in the crops soybean, potato or any other suitable plant).With regard to some plant, the number of times that increases from same rhizome results also is possible.The harvest cycle that changes plant may cause every acre year biomass yield increase (because the number of times of (for example in 1 year) any specified plant growth and results increases).The increase of growth velocity also may allow the geographic region cultivation transgenic plant wider than their wild type counterparts because the territorial restrictions of planting plant during usually by plantation when (season early) or results (season in evening) hostile environment condition determined.If the shortening harvest cycle can be avoided these unfavourable condition.Growth velocity can be determined by obtaining various parameters from growth curve drawing growth experiment, these parameters can be: T-Mid (plant reaches 50% required time of its largest amount) and T-90 (plant reaches 90% required time of its largest amount), or the like.

Implement the plant that method generation of the present invention has the growth velocity of increase.Therefore, according to the present invention, provide a kind of method that is used to increase the growth velocity of plant, this method is included in the nucleic acid of introducing in the plant and expressing coding S3a polypeptide.The increase of growth velocity is illustrated as respectively here for control plant/corresponding wild-type plant, and the seed weight that TKW increases, (full) seed number increases increases and harvest index increases.

Compare with control plant, plant be under the non-stress situation or plant contact multiple stress the time increase of productive rate and/or growth velocity takes place.Plant generally by more lentamente the growth reply contacted stress.In serious stress descending, plant even may stop growing fully.On the other hand, moderate stress be defined as plant contact in the text any can not cause that plant is dormant fully stress.Since the progress of the farming method (irrigation, fertilising, pesticide treatments), the crop plants of cultivation usually can not suffer from serious stress.Therefore, the impaired growth of moderate stress-induced usually is the characteristics that agricultural is not expected.Moderate stress be that the typical case that plant may contact stress.These stress be plant contact daily biology and/or abiotic (environment) stress.Typical abiotic or environmental stress comprises the thermal stress that heat or the cold/freezing temp by abnormality causes; Salt stress; Water is coerced (arid or excess water).Abiotic stress also may be that chemical causes.Biology stress generally be that those that caused by pathogenic agent such as bacterium, virus, fungi and insect stress.

Preferably can in any plant, change above-mentioned growth characteristics.

Term as used herein " plant " comprises whole plants, and ancestors of plant and offspring, and the part of plant comprise seed, branch, and stem, leaf, root (comprising stem tuber), and tissue and organ, wherein above-mentioned each all comprises goal gene.Term " plant " also comprises embryo, the meristematic tissue zone, and gametophyte, sporophyte, pollen and sporule, equally wherein above-mentioned each also all contains goal gene.

Especially the plant that can be used for the inventive method comprises all plants that belong to vegitabilia (Viridiplantae) superfamily, particularly unifacial leaf and dicotyledons, comprise feed or feed leguminous plants, ornamental plant, food crop, be selected from the arbor or the shrub of following species: Acacia species (Acaciaspp.), maple species (Acer spp.), Actinidia species (Actinidia spp.), Aesculus species (Aesculus spp.), New Zealand kauri (Agathis australis), Albiziaamara, Alsophila tricolor, Andropogon species (Andropogon spp.), Arachis species (Arachis spp.), betel nut (Areca catechu), Astelia fragrans, Astragaluscicer, Baikiaea plurijuga, Betula (Betula spp.), Btassica (Brassica spp.), Bruguiera conjugata (Bruguiera gymnorrhiza), Burkea africana, palas (Butea frondosa), Cadaba farinosa, Zhu Ying Pittosporum species (Calliandra spp.), daye tea (Camelliasinensis), Canna generalis Bailey (Canna indica), Capsicum species (Capsicum spp.), Cassia species (Cassia spp.), apart from pea (Centroema pubescens), Chaenomeles species (Chaenomeles spp.), Chinese cassia tree (Cinnamomum cassia), fruitlet coffee (Coffeaarabica), Colophospermum mopane, Coronillia varia, Cotoneaster serotina, hawthorn species (Crataegus spp.), Cucumis species (Cucumis spp.), Cupressus species (Cupressus spp.), Cyathea dealbata Quinces Quince (Cydonia oblonga), Japanese cypress (Cryptomeria japonica), Cymbopogon species (Cymbopogon spp.), Cyntheadealbata Quinces Quince (Cydonia oblonga), Dalbergia monetaria, Da Ye Rhizome of Fortune's Drynaria (Davallia divaricata), mountain horseleech species (Desmodium spp.), coarse freshwater mussel shellfish fern (Dicksonia squarosa), Diheteropogon amplectens, Dioclea spp, sickle Dolichos species (Dolichos spp.), Dorycnium rectum, Echinochloa pyramidalis, Ehrartia spp. Finger-millet (Eleusine coracana), Herba Eragrostidis pilosae species (Eragrestis spp.), Erythrina species (Erythrina spp.), eucalyptus species (Eucalyptus spp.), Eucleaschimperi, Eulalia villosa, Fagopyrum species (Fagopyrum spp.), feijoa (Feijoasellowiana), Fragaria species (Fragaria spp.), Moghania species (Flemingia spp), Freycinetia banksii, Geranium thunbergii, ginkgo (Ginkgo biloba), Glycinejavanica, Gliricidia spp, upland cotton (Gossypium hirsutum), Grevillea species (Grevillea spp.), Guibourtia coleosperma, rock Astragalus species (Hedysarumspp.), Hemarthria compressa (Hemarthia altissima), turn round Huang Mao (Heteropogon contortus), barley (Hordeum vulgare), Hyparrhenia rufa, Herba Hyperici Erecti (Hypericum erectum), Hyperthelia dissoluta, spend front yard indigo plant (Indigo incarnata) in vain, Jris species (Iris spp.), Leptarrhena pyrolifolia, lespedeza species (Lespediza spp.), Lettuca spp., Leucaena leucocephala, Loudetia simplex, sieve beans (Lotonus bainesii) that pause, Lotus species (Lotus spp.), Macrotyloma axillare, Malus species (Malusspp.), Manihot esculenta, alfalfa (Medicago sativa), metasequoia (Metasequoiaglyptostroboides), powder bajiao banana (Musa sapientum), Nicotiana species (Nicotianumspp.), donkey food Macroptilium species (Onobrychis spp.), Ornithopus spp., Oryza species (Oryzaspp.), Peltophorum africanum, Pennisetum species (Pennisetum spp.), Perseagratissima, green winter Solanum species (Petunia spp.), Phaseolus species (Phaseolus spp.), betel nut bamboo (Phoenix canariensis), Phormium cookianum, Photinia species (Photiniaspp.), white spruce (Picea glauca), Pinus species (Pinus spp.), pea (Pisumsativum), alpine totara (Podocarpus totara), Pogonarthria fleckii, Pogonarthria squarrosa, Populus species (Populus spp.), algarroba (Prosopiscineraria), Pseudotsuga menziesii (Mirbel) Franco (Pseudotsuga menziesii), Pterolobium stellatum, European pear (Pyrus communis), oak species (Quercus spp.), Rhaphiolepsisumbellata, delicious rod is spent palm fibre (Rhopalostylis sapida), Rhus natalensis, Europe gooseberry (Ribes grossularia), currant species (Ribes spp.), acacia (Robiniapseudoacacia), rose species (Rosa spp.), rubus species (Rubus spp.), Salix species (Salix spp.), Schyzachyrium sanguineum, parasol pine (Sciadopitysverticillata), sequoia sempervirens (Sequoia sempervirens), big tree (Sequoiadendrongiganteum), dichromatism chinese sorghum (Sorghum bicolor), spinach species (Spinacia spp.), Sporobolus fimbriatus, Stiburus alopecuroides, Stylosanthos humilis, Triquetrous Tadehagi Herb species (Tadehagi spp.), southern cypress (Taxodium distichum), Arabic Herba Themedae japonicae (Themeda triandra), Clover species (Trifolium spp.), Triticum species (Triticum spp.), tsuga heterophylla (Tsuga heterophylla), genus vaccinium species (Vacciniumspp.), Vetch species (Vicia spp.), grape (Vitis vinifera), the fertile gloomy flower (Watsonia pyramidata) of awl fringe, common calla (Zantedeschia aethiopica), corn (Zeamays), Amaranthus, choke, asparagus, cabbage, brassica oleracea var gemmifera, Caulis et Folium Brassicae capitatae, canola, Radix Dauci Sativae, Cauliflower, celery, the kale green plant, flax, kale; root of Szemao crotalaria; rape; gumbo; onion; potato; rice; soybean; strawberry; sugar beet; sugarcane; Sunflower Receptacle; tomato; pumpkin; tealeaves and algae, or the like.According to a preferred embodiment of the invention, plant is crop such as soybean, Sunflower Receptacle, rape, alfalfa, Semen Brassicae campestris, cotton, tomato, potato or tobacco.Still more preferably, plant is a monocotyledons, as sugarcane.More preferably, plant is a cereal, as rice, and corn, wheat, barley, grain, rye, Chinese sorghum or oat.

Term S3a is well known in the art.The title selected of the S3a that provides in the following table 1 from Naora and Naora (Immunology and Cell Biology (1999) 77,197-205).

Homologue and the name of table 1:S3a

Title	Species	Standard of perfection	Reference
				RPS3a	Human	Ribosomal protein	Metspalu etc., 1992
nbl	Human	The abundant expression in Namalwa Burkitt lymphoma cell	Naora etc., 1995
				RPS3a	Cat	The abundant expression in feline leukaemia virus inductive lymphoma	Starkey and Levy, 1995
fte-1	Rat	Destroyed in the inoblast that revertant v-fos transforms	Kho and Zarbl, 1992
				TU-11	Mouse	By the TNF-a abduction delivering	Gordon etc., 1992
13T	Mouse	The rat hybrid cell is abundant the expression in the heterozygote that transforms	Lecomte etc., 1997
				KRP-A	Sea hare	The abundant expression in neurocyte	Auclair etc., 1994
C3	Mosquito	In ovary, preferentially express	Zurita etc., 1997
				RPS3a	Fruit bat	Sudden change with the weak tendency phenotypic correlation	Van Beest etc., 1998
cyc07	Higher plant	The S phase is specific expressed	Ito etc., 1991
				MFT1	Yeast	Weakening protein is input to the sudden change in the plastosome	Garrett etc., 1991

By search sequence (preferred protein sequence) and known S3a protein sequence are compared, can identify S3a (for example referring to the sequence alignment shown in Fig. 1 and 2) at an easy rate.For example, (InforMax, Bethesda MD), adopt that the open point penalty in room is 10, to extend point penalty be 0.05 default setting in the room, and search sequence (with known S3a sequence) is compared to program can to use VNTI AlignX sequence multiple ratio.Because the S3a sequence is a high conservative, by those zones of any conservative region of search sequence and known S3a sequence are compared, those of ordinary skills can identify other S3a sequence at an easy rate.The zone of these high conservatives in Fig. 2 with 3 frame district graphic extensions.Therefore, the search sequence with corresponding conservative region will be accredited as S3a.Shown that also S3a combines (Westermann etc. (FEBSLetters Vol.97, No.1 (in January, 1979)) with eukaryotic initiation factor eIF-2 and eIF-3.

Term as used herein " S3a " is used in reference to protein, when it is used for sequence alignment [for example comparison shown in Fig. 1 or 2], is with the comparison of known S3a protein sequence and comprises and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.The S3a coding nucleic acid of mentioning in the literary composition is meant the proteinic nucleic acid that coding is such, when it is used for sequence alignment [for example comparison shown in Fig. 1 or 2], is with the comparison of known S3a protein sequence and comprises and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.S3a and S3a coding nucleic acid are useful in the method for the invention as defined above.

Nucleic acid in plant to be imported can be any as defined above S3a encoding sequence.Nucleic acid is preferably shown in SEQ ID NO:1.Nucleic acid in plant to be imported preferably effectively is connected to cross in plant with constitutive promoter expresses.Constitutive promoter is the GOS2 promotor preferably, more preferably from the GOS2 promotor of rice.Obviously, application of the present invention is not limited to the S3a shown in the SEQ ID NO:1, and when by the GOS2 promoters driven, application of the present invention also is not limited to S3a gene/expression of nucleic acids.

According to a preferred aspect of the present invention, consider the S3a expression of nucleic acids of enhanced or increase.Enhanced or the gene of increase or the method for gene product expression of obtaining fully reported in this area, and for example, it comprises by crossing of strong promoter driving expresses the use of transcriptional enhancer or translational enhancer.

The nucleic acid of coding S3a can derive from any source.Nucleic acid/gene of coding S3a can separate from microbe-derived, as bacterium, yeast or fungi, or separates and originates from plant, algae or animal (comprising the people).Can the natural form that be in the nucleic acid in composition and/or the genome environment be modified by deliberate manual operation.Nucleic acid is homologous nucleic acid preferably, can be will be imported into wherein identical plant species from it available from the nucleic acid of plant promptly, also can be from different plant species.Nucleic acid can separate from dicotyledonous kind, and preferably from Cruciferae, also preferable separation is from Arabidopis thaliana (Arabidopsis thaliana).More preferably, separate S3a from Arabidopis thaliana shown in SEQ ID NO:1, its aminoacid sequence is shown in SEQ ID NO:2.

Sequence description shown in the SEQ ID NO:1 from the S3a of Arabidopis thaliana, SEQ ID NO:2 is an amino acid sequence corresponding.Best, application of the present invention is not limited to the S3a from Arabidopis thaliana of use shown in SEQ ID NO:1.Also can use any S3a or the S3a encoding sequence of definition as mentioned to implement method of the present invention.

The S3a or the S3a coding nucleic acid that are used to implement the inventive method can be:

(i) part of S3a coding nucleic acid, the preferably part of the S3a coding nucleic acid shown in SEQ ID NO:1;

(ii) can with the sequence of S3a coding nucleic acid hybridization, preferably can with the sequence of S3a coding nucleic acid hybridization shown in SEQ ID NO:1;

The (iii) splice variant selected of S3a coding nucleic acid, the preferably splice variant selected of the S3a coding nucleic acid shown in SEQ ID NO:1;

The (iv) allelic variant of S3a coding nucleic acid, the preferably allelic variant of the S3a coding nucleic acid shown in SEQ ID NO:1; With

(v) S3a albumen/amino acid whose homologue and derivative, preferably proteic homologue of S3a and the derivative shown in SEQ ID NO:2.

Above-mentioned part, hybridization sequences, splice variant and allelic variant, homologue and derivative are S3a or in the range of definition of S3a coding nucleic acid those that fall into as hereinbefore defined, promptly, under proteinic situation, with the comparison of known S3a protein sequence and comprise and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment; And under the situation of nucleic acid, coding is compared with known S3a protein sequence and is comprised and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.

It is evident that for those of ordinary skills, use the S3a dna sequence dna of total length, is not to be necessary for implementing method of the present invention.Method of the present invention is preferably used for example part of the DNA/ nucleic acid of the coding S3a shown in the SEQ ID NO:1.Partly refer to derive from primary (bigger) dna molecular or section of DNA prepared therefrom.For example, can use technology well known in the art, carry out one or more disappearances by nucleotide sequence and prepare described part SEQ ID NO:1.The part that is suitable for implementing the inventive method is proteic those parts of coding S3a, and this S3a albumen is compared with known S3a protein sequence and comprised and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.

Therefore according to the present invention, a kind of method that is used to improve the growth characteristics of plant is provided, be included in the part of introducing and express the S3a coding nucleic acid in the plant, preferably the part of the nucleic acid shown in SEQ ID NO:1.

Also can be used for the method for carrying out an invention be can with the nucleic acid of S3a coding nucleic acid hybridization, preferably can with the nucleic acid of S3a nucleic acid sequence encoding hybridization shown in SEQ ID NO:1.Such hybridization sequences is proteic those sequences of coding S3a, and this S3a protein sequence and known S3a protein sequence are compared and comprised and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.

Term " hybridization " is a homologous complementary nucleotide sequence annealed process each other basically wherein as herein defined.Crossover process can occur in the solution fully, and promptly two complementary nucleic acid all are in the solution.The biology tool that relies on this method comprises polymerase chain reaction (PCR; And based on this all methods), synthetic, the RNA difference of subtractive hybridization, random primer extension, S1 nuclease mapping, primer extension, reverse transcription, cDNA shows and dna sequencing.Crossover process can also so be carried out, and promptly wherein one of complementary nucleic acid is fixed on matrix such as magnetic bead, sepharose pearl or any other resin.The biology tool that relies on this method comprises the separation of poly (A+) mRNA.Crossover process can so be carried out in addition, promptly wherein one of complementary nucleic acid is fixed on solid support such as nitrocellulose membrane or the nylon membrane, or for example be fixed on the silicate glasses upholder (latter is referred to as nucleic acid array or microarray, or is referred to as nucleic acid chip) by for example photolithography.The biology tool that relies on this method comprises RNA and dna gel engram analysis, colony hybridization, plaque hybridization, in situ hybridization and microarray hybridization.In order to make hybridization take place, make nucleic acid molecule thermally denature or chemical modification usually, so that two strands is unwind into two strands and/or removed hair clip or other secondary structure from single-chain nucleic acid.The preciseness of hybridization is subjected to such as condition effect such as temperature, salt concn and hybridization buffer compositions.Hybridization is preferably carried out under rigorous condition.Rigorous condition adopts low ionic strength and high-temperature for (1) and washs, for example, and 0.5M sodium phosphate buffer pH 7.2, be dissolved in the 1mM EDTA pH 8.0 of 7%SDS, temperature is 65 ℃ or 55 ℃, and perhaps (2) use denaturing agent such as methane amide during hybridizing, for example, 50% (volume/volume) methane amide adds 0.1% bovine serum albumin, 0.1%Ficoll, 0.1% polyvinylpyrrolidone, 0.05M sodium phosphate buffer pH 6.5 adds 0.75MNaCl, 0.075M Trisodium Citrate, temperature are 42 ℃.Concrete example comprises use 50% methane amide, 5 * SSC (0.75 NaCl, 0.075M Trisodium Citrate), 50mM sodium phosphate (pH 6.8), 0.1% trisodium phosphate, 5 * Denhard ' s liquid, the salmon sperm DNA of supersound process (50nm/ml), 0.1%SDS and 10% dextran sulfate, temperature are 55 ℃, and wash in 0.2 * SSC and 0.1%SDS in 55 ℃.Those skilled in the art can determine at an easy rate and suitably change rigorous condition, to obtain clear and detectable hybridization signal.

Therefore according to the present invention, a kind of method that is used to improve the growth characteristics of plant is provided, be included in introduce in the plant and express preferably under rigorous condition can with the sequence of S3a nucleic acid sequence encoding [preferably with shown in SEQ ID NO:1 S3a coding nucleic acid] hybridization.

The splice variant selected of S3a coding nucleic acid, preferably the variant selected of the S3a coding nucleic acid shown in SEQ ID NO:1 also can be used for implementing method of the present invention.The splice variant that is suitable for implementing the inventive method is proteic those splice variants of coding S3a, and this S3a albumen is compared with known S3a protein sequence and comprised and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.Such nucleotide sequence variant contained in term " selectable splice variant " as used herein, and promptly wherein selected intron and/or exon excise, replace or add.Such variant is that wherein proteinic biologic activity keeps unaffected those variants, and it can obtain by the function fragment of retaining protein optionally.Such splice variant can be natural discovery maybe can be synthetical.The method that is used to prepare such splice variant is well known in the art.

Therefore, the present invention also provides a kind of method that is used to improve the growth characteristics of plant, is included in the splice variant selected of introducing and expressing the S3a coding nucleic acid in the plant, preferably the splice variant selected of the S3a coding nucleic acid shown in SEQ ID NO:1.

The allelic variant of S3a coding nucleic acid, preferably the allelic variant of the S3a coding nucleic acid shown in SEQ ID NO:1 also can be used for implementing method of the present invention.The allelic variant that is suitable for implementing the inventive method is proteic those allelic variants of coding S3a, and this S3a albumen is compared with known S3a protein sequence and comprised and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.

The natural existence of allelic variant, and to include be these natural allelic purposes within method of the present invention.Allelic variant comprises single nucleotide polymorphism (SNP), and little insertion/deletion polymorphism (INDEL).The size of INDEL is usually less than 100bp.In the naturally occurring polymorphic bacterial strain of most of organisms, SNP and INDEL have constituted a maximum class sequence variants.

Therefore, the present invention also provides a kind of method that is used to improve the growth characteristics of plant, is included in the allelic variant of introducing and expressing the S3a coding nucleic acid in the plant, preferably the allelic variant of the S3a coding nucleic acid shown in SEQ ID NO:1.

In addition, method of the present invention preferably also can be used homologue, derivative or the active fragments of the S3a of S3a shown in SEQ ID NO:2.Use routine techniques well known to those skilled in the art, can determine the homologue of coding S3a or the nucleic acid of derivative at an easy rate.

Proteinic " homologue " contain with respect to the unmodified protein matter of being discussed have aminoacid replacement, disappearance and/or insertion and to its derived from unmodified protein matter have peptide, oligopeptides, polypeptide, protein and the enzyme of similar biologic activity and functionally active.In order to produce such homologue, other amino acid that proteinic amino acid can be had similar performance (as similar hydrophobicity, wetting ability, antigenicity, form or interrupt the trend of α-Luo Xuanjiegou or β-laminated structure) replaces.The conservative substitution table is (for example referring to Creighton (1984) Proteins.W.H.Freeman and Company) well known in the art.

At least 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% the sequence identity that can be used for that the homologue of the inventive method and the aminoacid sequence shown in the SEQ ID NO:2 have that preference increases progressively successively.And the homologue that is suitable for implementing the inventive method is with known S3a protein sequence comparison and comprises those homologues with the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.

The homology of two kinds of special shapes also contained in term " homologue ", and it comprises directly that to homology (orthologous) sequence and symbiosis homology (paralogous) sequence they have contained the evolution notion that is used to describe the gene genetic relation.Term " symbiosis homology " relates to the gene replication in certain species genome, produces paralogous gene.Term " directly to homology " relates in the different organisms because of the homologous gene due to the genetic affinity.

For example, directly can find at an easy rate by implementing so-called mutual blast search in the monocotyledons species to homologue.This can finish by the blast first time, be included in any sequence library as in public's available ncbi database that can in http://www.ncbi.nlm.nih.gov, find, the sequence of being discussed (for example, SEQ ID NO:1 or SEQ ID NO:2) is carried out blast.If seek be rice directly to homologue, the sequence of being discussed will be carried out blast with 28,469 the full length cDNA clone of for example going up available Oryza sativa Nipponbare rice from NCBI.When Nucleotide begins, using BLASTn,, and use standard default (expected value 10, comparison value 50) maybe when when protein begins, can using TBLASTX.The result of blast can filter.And then with filter result or not the full length sequence of filter result carry out reverse blast (blast for the second time) with the sequence (SEQ IDNO:1 or 2) discussed.Then the result of blast for the first time and is for the second time compared.Under the situation of extended familys, use ClustalW, then use adjacent threaded tree to help to observe cluster (clustering).

Homologue can be the form of protein " replacement variant ", that is, wherein at least one residue in the aminoacid sequence is removed, and a different residue is inserted on its position.Aminoacid replacement generally is single residue, but depends on that the function restriction that polypeptide is constituted can be a bunchiness; Insert normally about 1 to 10 amino-acid residue, lack and be about 1 to 20 residue.Preferably, aminoacid replacement comprises that conserved amino acid replaces.

Homologue can also be proteinic " insertion variant ", that is, wherein one or more amino-acid residues are introduced in the predetermined site in the protein.Insertion can comprise that N-terminal and/or C-terminal merge, and inserts in the single or multiple amino acid whose sequence.Usually, the insertion in the aminoacid sequence will be merged less than amino or C-terminal, be about the order of magnitude of 1 to 10 residue.The example of amino or C-terminal fused protein or peptide comprises binding domains or activation structure territory, bacteriophage coat protein, (Histidine) 6-label, glutathione S-transferase-label, A albumen, maltose binding protein, Tetrahydrofolate dehydrogenase, Tag100 epi-position, c-myc epi-position, FLAG  epi-position, lacZ, CMP (calmodulin binding peptide), HA epi-position, C albumen epi-position and the VSV epi-position as the activating transcription factor that is used for the two heterological systems of yeast.

The homologue of protein " disappearance variant " form is characterised in that from protein removes one or more amino acid.

Use peptide synthetic technology well known in the art, synthesize or the like, or, can prepare proteinic amino acid variant at an easy rate by the recombinant DNA operation as solid-phase peptide.The method that dna sequence dna is operated to produce proteinic replacement, insertion or disappearance variant is well known in the art.For example, it is well known to those skilled in the art producing the technology that replaces sudden change at the predetermined site of DNA, comprise M13 mutagenesis, T7-Gen vitro mutagenesis (USB, Cleveland, OH), the site-directed mutagenesis (Stratagene of Fast transforms, San Diego, CA), site-directed mutagenesis or other site-directed mutagenesis scheme of PCR-mediation.

" derivative " comprises peptide, oligopeptides, polypeptide, protein and enzyme, compares with the aminoacid sequence of the natural existence form of S3a albumen, and for example, shown in SEQ ID NO:2, it can comprise that there be replacement, disappearance or the interpolation of amino-acid residue in natural and non-natural.Proteinic " derivative " comprises peptide, oligopeptides, polypeptide, protein and enzyme; compare with the aminoacid sequence of the natural existence form of polypeptide, it can comprise the naturally occurring amino-acid residue of change, glycosylated, acidylate or the amino-acid residue that non-natural exists.With its derived from aminoacid sequence compare, derivative also can comprise one or more non-aminoacid replacement bases, for example with aminoacid sequence covalently or non-covalently bonded reporter molecules or other part, as in conjunction with so that the reporter molecule of its detection, and with respect to the natural amino-acid residue that exists the non-natural for the proteinic aminoacid sequence to exist.

The method of search and evaluation S3a homologue is well known to those skilled in the art.The method that the sequence that is used for comparison is compared is well known in the art, and these class methods comprise GAP, BESTFIT, BLAST, FASTA and TFASTA.GAP uses the algorithm of Needleman and Wunsch (J.Mol.Biol.48:443-453,1970) to realize the comparison of two complete sequences, the number of its maximization coupling, and minimize the room number.BLAST arithmetic calculation sequence identity per-cent, and similarity between two sequences carried out statistical analysis.The software of carrying out the BLAST analysis passes through NCBI (the National Centre for Biotechnology Information) but public's acquisition.Can be by getting the full length protein sequence, use the multiple contrast program of VNTI AlignX sequence that they are carried out sequence alignment (InforMax, Bethesda, MD), and use the open point penalty in room be 10 and the room extend to 0.05 default settings, identify the homologue that is suitable for the inventive method, that is, and those homologues that have at least 55% sequence identity with the aminoacid sequence shown in the SEQ ID NO:2.For example referring to Fig. 1.

Therefore, the present invention also provides a kind of method that is used to improve the growth characteristics of plant, be included in and introduce and express the homologue of the S3a of coding shown in SEQ ID NO:2 or the nucleic acid of derivative in the plant, this homologue, aminoacid sequence shown in derivative or active fragments and the SEQ ID NO:2 have that preference increases progressively successively at least 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95% sequence identity, and this homologue or derivative and the comparison of known S3a protein sequence and comprise and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.

The present invention also provides genetic constructs and carrier, so that introduce and/or express the nucleotide sequence that can be used for the inventive method.

Therefore, the invention provides a kind of gene construct, it comprises:

(i) S3a coding nucleic acid is preferably shown in SEQ ID NO:1;

(ii) can drive one or more control sequences that (i) amplifying nucleic acid sequence is expressed; Randomly

(iii) transcription termination sequence.

The construct that can be used for the inventive method can use recombinant DNA technology well known to those skilled in the art to make up.Gene construct can be inserted in the carrier, and this carrier can be purchased, and is suitable for being transformed in the plant, and is suitable for expressing goal gene in cell transformed.

Plant transforms with the gene construct that comprises aim sequence (i.e. S3a coding nucleic acid as hereinbefore defined).Aim sequence is connected effectively with one or more control sequences (at least with a promotor).Term " controlling element ", " control sequence " and " promotor " can be exchanged use in the text, and should get its broad sense, are meant the regulation and control nucleotide sequence that can work to the expression of the sequence that is connected with them.Above-mentioned term is contained and is derived from classical eukaryotic gene group gene and (comprise the TATA box that accurate transcription initiation is required, with or without CCAAT box sequence) and according to growing and/or external stimulus, or change the transcription regulating nucleotide sequence of the other controlling element (that is, upstream activating sequence, enhanser and silencer) of genetic expression in the tissue specificity mode.This term also comprises the transcription regulating nucleotide sequence of classical prokaryotic gene, and in this case, it may comprise-35 box sequences and/or-10 box transcription regulating nucleotide sequences.Synthetic fusion molecule or derivative also contained in term " controlling element ", and it makes nucleic acid molecule can, express in cell, tissue or organ, activates or strengthen this expression.Term " effectively connect " refers to functional connection the between promoter sequence and the goal gene as used herein, so that promoter sequence can start transcribing of goal gene.

Best, the promotor of any kind all can be used for driving the expression of S3a coding nucleic acid.

Preferably, the nucleic acid of coding S3a is connected with constitutive promoter effectively.Refer to this promotor predominant expression in more than a kind of tissue or organ as defined term " composing type " in the literary composition, and at any stage of plant life cycle predominant expression all.Preferably, constitutive promoter predominant expression in whole strain plant.Preferably, constitutive promoter is the GOS2 promotor from rice.

The example that is suitable for other constitutive promoter of the inventive method A that is listed in the table below.

Table A: the example that is used to implement constitutive promoter of the present invention

Gene source	Expression pattern	Reference
			Actin muscle	Composing type	McElroy etc., Plant Cell, 2:163-171,1990

CAMV 35S	Composing type	Odell etc., Nature, 313:810-812,1985
			CaMV 19S	Composing type	Nilsson etc., Physiol.Plant.100:456-462,1997
GOS2	Composing type	De Pater etc., Plant J Nov; 2 (6): 837-44,1992
			Ubiqutin	Composing type	Christensen etc., Plant Mol.Biol.18:675-689,1992
The rice cyclophilin	Composing type	Buchholz etc., Plant Mol Biol.25 (5): 837-43,1994
			Corn H3 histone	Composing type	Lepetit etc., Mol.Gen.Genet.231:276-285,1992
Actin muscle 2	Composing type	An etc., Plant are (1) J.10; 107-121,1996

Randomly, one or more terminator sequences also can be used for being introduced in the construct in the plant.Control sequence contained in term " terminator ", and it is the dna sequence dna that is positioned at the transcriptional units end, transmits the signal of 3 ' processing and primary transcript polyadenylation and Transcription Termination.Other controlling element also can comprise transcriptional enhancer and translational enhancer.Those skilled in the art know and are suitable for implementing terminator of the present invention and enhanser.Such sequence is known, or those skilled in the art can obtain at an easy rate.

Genetic constructs of the present invention can also be included in keeps and/or duplicates required replication origin sequence in the particular cell types.An example is the situation that needs genetic constructs to keep as extrachromosomal inheritance element (for example, plasmid or clay molecule) in bacterial cell.Preferred replication origin is including, but not limited to f1-ori and colE1.

Genetic constructs randomly can comprise selectable marker gene.As used herein, term " selectable marker gene " comprises gives the cell phenotype of expressing this gene so that identify and/or select any gene with nucleic acid construct transfection of the present invention or cell transformed.Suitable mark can be selected from the mark of giving microbiotic or Herbicid resistant, and it is introduced new metabolic proterties or allows vision to select.The example of selectable marker gene comprises can give antibiotics resistance (as make the nptII of Xin Meisu and kantlex phosphorylation, or make the hpt of Totomycin phosphorylation), Herbicid resistant (for example provides the bar to the resistance of Basta; AroA or gox to the resistance of glyphosate are provided) gene, or provide the gene (as allowing the manA of plant utilization seminose) of metabolic proterties as sole carbon source.Visual marker gene cause forming color (beta-glucuronidase for example, GUS), luminous (as luciferase) or fluorescence (green fluorescent protein GFP and derivative thereof).

The present invention is also contained can be by the plant of method acquisition of the present invention.Therefore the present invention provides and can introduce and express the proteic nucleic acid of coding S3a as hereinbefore defined in this plant by the plant of method acquisition of the present invention.

The present invention also provides the method for the transgenic plant of the growth characteristics that a kind of production has improvement, is included in to introduce and express S3a coding nucleic acid, the preferably nucleic acid shown in SEQ ID NO:1 in the plant.

More particularly, the invention provides the method for transgenic plant that a kind of production has the growth characteristics of improvement, this method comprises:

(i) in plant or vegetable cell, introduce S3a coding nucleic acid, the preferably nucleic acid shown in SEQ ID NO:1;

(ii) promoting regeneration and making culturing plants cell under the sophisticated condition of plant-growth.

Nucleic acid can be introduced directly among vegetable cell or the plant itself (comprising the tissue, organ or any other parts that are incorporated into plant).According to a preferable feature of the present invention, nucleic acid preferably is introduced in the plant by conversion.

The term mentioned as this paper " conversion " contained the transfer of exogenous polynucleotide in host cell, and no matter the method that shifts how.Can vegetative subsequently plant tissue, no matter be to take place by organ or embryo, can transform with genetic constructs of the present invention, and from its complete plant that regenerates.The particular organization of selecting will look the vegetative propagation system of the specific species that can be used for and be suitable for transforming most and change.Exemplary target tissue (for example comprises leaf disc, pollen, embryo, cotyledon, hypocotyl, megagametophyte, callus, already present meristematic tissue, apical meristem, bud and root meristematic tissue that armpit is given birth to) and inductive meristematic tissue (for example, cotyledon meristematic tissue and hypocotyl meristematic tissue).Polynucleotide can instantaneous or stably be introduced in the host cell, and can keep the nonconformity state, for example, and as plasmid.Selectively, it can be incorporated in the host genome.The transformed plant cells of gained can be used for being regenerated as the conversion plant with the method known to those skilled in the art then.

The conversion of plant species is a kind of quite conventional technology at present.Advantageously, any method for transformation all can be used for introducing goal gene in suitable progenitor.Method for transformation comprise use liposome, electroporation, can increase chemical that dissociative DNA takes in, direct injection DNA in plant, the particle gun partickle bombardment, use virus or pollen transforms and microinjection.Method optional from be used for protoplastis calcium/polyoxyethylene glycol method (Krens, F.A. etc., 1882, Nature 296,72-74; Negrutiu I. etc., June 1987, Plant Mol.Biol.8,363-373); The electroporation of protoplastis (Shillito R.D. etc., 1985 Bio/Technol 3,1099-1102); Microinjection in vegetable material (Crossway A. etc., 1986, Mol.Gen Genet 202,179-185); DNA or the bombardment of RNA coating particles (Klein T.M. etc., 1987, Nature 327,70); With (nonconformity type) virus infection or the like.The transgenosis rice plant of expressing S3a preferably uses any well-known process that rice transforms that is used for, conversion by soil Agrobacterium (Agrobacterium) mediation produces, how the method for describing in the document of playing for example in office: disclosed European patent application EP 1198985 A1, Aldemita and Hodges (Planta, 199,612-617,1996); Chan etc. (Plant Mol.Biol.22 (3) 491-506,1993), Hiei etc. (Plant is (2) 271-282 J.6,1994), its disclosed content is incorporated in the literary composition as a reference as the full content of its statement.Transform (Nat.Biotechnol.1996 Jun such as preferable methods such as Ishida as for cereal; 14 (6): 745-50) or (Plant Physiol.2002 May such as Frame; 129 (1): 13-22), its disclosed content is incorporated in the literary composition as a reference as the full content of its statement.

Usually after transforming, the coded one or more marks of expressive gene of plant that move with the goal gene corotation in selection vegetable cell or the cell mass are followed the material regeneration that will transform and are become whole plant.

After DNA transfer and the regeneration, can estimate and infer existence, copy number and/or the genome structure that transforms goal gene in the plant, for example use Southern to analyze.Selectively or additionally, the expression level of newly introducing DNA can use Northern and/or Western to analyze and monitor, and two kinds of technology all are well known to those of ordinary skill in the art.

The conversion plant that produces can breed by the whole bag of tricks, as by vegetative propagation or classical breeding technique.For example, the first-generation (or T1) transforms the s-generation (or T2) transformant that plant can selfing be isozygotied with generation, and the T2 plant can further breed by the breeding technique of classics.

The inverting biological body that produces can be various forms of.For example, they can be the mosaics of transformant and non-transformed cell; Clone's transformant (for example, all cell is all transformed to contain expression cassette); Transform with the graft of unconverted tissue (for example, in plant, transform rhizome by grafting in unconverted scion).

Any vegetable cell or plant that the present invention obviously prolongs and produced by any method of describing in the literary composition, and all plant parts and propagulum thereof.The present invention also prolongs and includes the former generation conversion that produced by any above-mentioned method or the offspring of transfectional cell, tissue, organ or complete plant, and unique requirement is that this offspring shows genotype and/or the phenotypic characteristic identical with genotype that is produced by method of the present invention and/or phenotypic characteristic in parental generation.The present invention also comprises the host cell that contains separative S3a coding nucleic acid.Preferred host cell is a vegetable cell according to the present invention.The present invention also prolongs and the part gathered in the crops of plant, as but be not limited to seed, leaf, fruit, flower, stem culture, rhizome, stem tuber and bulb.

Also can implement method of the present invention and not need in plant to introduce the nucleic acid of coding S3a.This can realize (preferably in the locus of S3a encoding gene) by introducing genetic modification.Locus as defined gene in the literary composition is meant genome area, and it comprises the upstream or the downstream of goal gene and 10KB coding region.

For example, genetic modification can be introduced by following any (or multiple) method: TDNA activation, TILLING, site-directed mutagenesis, homologous recombination, directly evolution, or as indicated above, by in plant (cell), introducing and express the S3a coding nucleic acid.

T-DNA activates the insertion that label Science (1992) 1350-1353 such as () Hayashi comprises T-DNA, it contains the promotor (also may be translational enhancer or intron) of structure like this usually in the upstream of the genome area of goal gene or gene coding region or downstream 10KB place, thereby this promotor can instruct target gene expression.Usually, destroy the expression regulation of target gene natural promoter, and this gene is under the control of promotor of new introducing it.Promotor generally is embedded among the T-DNA.For example, T-DNA infects by soil Agrobacterium and is inserted in the Plant Genome at random, and causes the gene overexpression near the T-DNA of this insertion.Because near the gene overexpression of the promotor of introducing, the transgenic plant of gained demonstrate the dominant phenotype.Promotor to be introduced can be any promotor that can instruct gene to express in required organism [is plant in this case].For example, composing type, organize preference, the cell type preference and inducible promoter all is suitable for T-DNA and activates.

Also can use TILLING technology (targeted induction genome local mutating technology), genetic modification is incorporated in the locus of nucleic acid/gene of coding S3a.This is a kind of induced-mutation technique, can be used for producing and/or identify, and final separation energy show the mutagenesis variant of the S3a coding nucleic acid of S3a biologic activity.TILLING can also select to carry the plant of this class mutation variants.These mutation variants even can show the higher S3a activity that natural form showed than this gene.TILLNG combines high-density mutagenesis and high-throughput screening method.The step that TILLING generally follows is: (a) EMS mutagenesis (Redei and Koncz, 1992; Feldmann etc., 1994; Lightner and Caspar, 1998); (b) DNA preparation and individual the merging; (c) pcr amplification in purpose zone; (d) sex change and annealing are to allow to form heteroduplex; (e) DHPLC, wherein the existence of heteroduplex detection is extra peak value in the color atlas in the storehouse; (f) identify mutated individual; (g) order-checking of sudden change PCR product.The method of TILLING is (McCallum Nat Biotechnol.2000 Apr well known in the art; 18 (4): 455-7, by Stemple 2004 (TILLING-a high-throughput harvest for functionalgenomics.Nat Rev Genet.2004 February; 5 (2): 145-50.) summary).

Can utilize site-directed mutagenesis to produce variant or its part of S3a coding nucleic acid.Several Methods can be used to realize site-directed mutagenesis, modal method (the current protocols inmolecular biology.Wiley volume that is based on PCR Http:// www.4ulr.com/products/currentprotocols / index.html)

Directly evolution also can be used for producing the variant of S3a coding nucleic acid.This comprises multiple DNA reorganization, then carries out suitable screening and/or selection to produce variant (Castle etc., (2004) Science 304 (5674): 1151-4 of S3a coding nucleic acid; United States Patent (USP) 5,811,238 and 6,395,547).

T-DNA activation, TILLING, site-directed mutagenesis and direct the evolution are the examples that can produce the technology of new allelotrope and SYT variant.

Homologous recombination can be introduced selected nucleic acid at genomic regulation selected location place.Homologous recombination is a normally used standard technique in the biology, is used for low organism such as yeast or the liver moss physcomitrella that waits.The method that is used for carrying out homologous recombination plant describes in model plant not only that (Extrachromosomal homologous recombination and genetargeting in plant cells after Agrobacterium-mediated transformation.1990EMBO such as Offringa is year October J.1990; 9 (10): 3077-84), and (Terada R is for example described in the rice cereal crop, Urawa H, Inagaki Y, Tsugane K, Iida S.Efficient gene targetingby homologous recombination in rice.Nat Biotechnol.2002.Iida and Terada:A tale of two integrations, transgene and T-DNA:gene targeting byhomologous recombination in rice.Curr Opin Biotechnol.2004 April; 15 (2): 132-8).For example, treat that the nucleic acid (it can be a S3a coding nucleic acid as hereinbefore defined) of target need not be targeted in the locus of S3a encoding gene, but be directed into the zone of high expression level.The nucleic acid for the treatment of target can be the allelotrope of improvement, is used to replace native gene, or additionally is incorporated in the native gene.

The present invention also comprises the purposes of nucleic acid of the S3a that encodes and the purposes of S3a polypeptide.

A kind of such purposes relates to as hereinbefore defined S3a certainly in the growth characteristics of improvement plant, especially for improving productive rate, the especially purposes of seed productive rate.The seed productive rate can comprise following one or more aspect: (full) seed number of increase, the seed weight of increase, the harvest index of increase and TKW of increase or the like.The S3a coding nucleic acid can be shown in SEQ ID NO:1, and S3a albumen can be the amino acid shown in SEQ ID NO:2.

The nucleic acid of coding S3a as hereinbefore defined and S3a polypeptide also have purposes in breeding program greatly.The S3a coding nucleic acid can be shown in SEQ ID NO:1, or S3a can be the amino acid shown in SEQ ID NO:2.For example, S3a coding nucleic acid or its part can be positioned on the karyomit(e) (or its part), and preferably relevant with one or more family member links together.In an example of this breeding system, evaluation may with the chain dna marker of gene genetic that can regulate in the plant the proteic expression of nucleic acid of coding S3a, this gene can be the gene of coding S3a albumen itself, or can influence active any other gene of coding proteic expression of gene of S3a and/or S3a albumen itself directly or indirectly.This then dna marker can be used for breeding system, has the plant of the growth characteristics of improvement with selection.

The allelic variant of S3a also can be used for conventional breeding system, as in the auxiliary breeding of mark.Such breeding system needs the mutagenic treatment by plant sometimes, introduces allelic variation in plant.A kind of suitable mutafacient system is EMS mutagenesis.Carry out the evaluation of allelic variant by for example PCR then.Then be the screening step, be used to select the good allelic variant of the sequence of discussing, it produces the growth characteristics of improvement in plant.Generally the growth characteristics that contain the plant of the different allelic variants (for example, the different allelic variants of SEQ ID NO:1) that sequence is discussed to some extent by monitoring are screened.The monitoring growth characteristic can be in the greenhouse or the field carry out.Optional step comprises and makes plant and another kind of plant hybridization in addition, wherein identifies good allelic variant.For example, this can be used for producing the purpose phenotypic characteristic of combination.

The S3a coding nucleic acid also can be used as probe, and so that gene is carried out heredity and physical mapping, these genes are a part and signs of chain with it proterties.These information can be used for plant breeding, have the kind system of desired phenotype with exploitation.This application of S3a coding nucleic acid, only needing length is the nucleotide sequence of at least 15 Nucleotide.The S3a coding nucleic acid can be used as restriction fragment length polymorphism (RFLP) mark.The Southern trace (Maniatis) of the plant genome DNA of restrictive diges-tion can use the S3a coding nucleic acid to survey.Can use a computer then program such as MapMaker (Lander etc. (1987) Genomics 1:174-181) carries out genetic analysis to the banding pattern of gained, to make up genetic map.In addition, this nucleic acid can be used for surveying the Southern trace, and this Southern trace contains the genomic dna that the restriction endonuclease of one group of individuality representing parent and clear and definite genetic cross offspring is handled.Record the separation of dna polymorphism, and be used for calculating the position (Botstein etc. (1980) Am.J.Hum.Genet.32:314-331) that the S3a coding nucleic acid uses the genetic map of this colony's acquisition in front.

The preparation and the purposes in genetic mapping of the probe in plant gene source are described among Bematzky and Tanksley (1986) the Plant Mol.Biol.Reporter 4:37-41.Numerous publications were described and are used method recited above or its flexible form that specific cDNA clone is carried out genetic mapping.For example, F2 hybridization colony, backcross population, panmictic population, the homogenic system of close relative and other group of individuals can be used for mapping.These class methods are well known to those skilled in the art.

Nucleic acid probe also can be used for physical mapping and (that is, sequence is placed on the physical map; Referring to Hoheisel etc.: Non-mammalian Genomic Analysis:A Practical Guide, Academicpress 1996, the 319-346 pages or leaves, and the reference of wherein quoting).

In another embodiment, nucleic acid probe can be used for direct fluorescence in situ hybridization (FISH) mapping (Trask (1991) Trends Genet.7:149-154).Though present FISH drawing method is partial to use big clone (several to hundreds of KB; Referring to (1995) Genome Res.5:13-20 such as Laan), but the raising of sensitivity allows to use short probe to carry out the FISH mapping.

Many various methods based on nucleic acid amplification that are used for heredity and physical mapping can use described nucleic acid to implement.Example comprises the polymorphism (CAPS of allele specific amplification (Kazazian (1989) J.Lab.Clin.Med11:95-96), pcr amplified fragment; Sheffield etc. (1993) Genomics16:325-332), allele-specific connects (Landegren etc. (1988) Science 241:1077-1080), Nucleotide extension (Sokolov (1990) Nucleic Acid Res.18:3671), radiation hybridization mapping (Walter etc. (1997) Nat.Genet.7:22-28) and Happy mapping (Dear and Cook (1989) Nucleic Acid Res.17:6795-6807).For implementing these methods, use nucleotide sequence design and preparation primer right, to be used for amplified reaction or primer extension reaction.This class primer design is well-known to those skilled in the art.In the method that adopts the PCR-based genetic mapping, has the difference that dna sequence dna between the parental generation of mapping corresponding to nucleotide sequence of the present invention zone is crossed in necessary evaluation.Yet this is dispensable usually to drawing method.

Nucleic acid and the S3a polypeptide of coding S3a also can be used as growth regulator.The S3a coding nucleic acid can be the nucleic acid shown in SEQ ID NO:1, and S3a albumen/polypeptide can be the amino acid shown in SEQ ID NO:2.Because these S3a can be used for improveing the growth characteristics of plant, S3a also is useful growth regulator, as weedicide or growth stimulator.Therefore the present invention provides a kind of composition, and it comprises the nucleic acid of S3a or coding S3a, together with suitable carriers, thinner or vehicle, as growth regulator.

Method of the present invention produces the plant of the growth characteristics with improvement, as previously discussed.The growth characteristics of these improvement also can combine with favourable proterties on other economics, as other volume increase proterties, to various stress tolerance, the proterties of modifying various constitutional featuress and/or biochemistry and/or physiologic character.

Description of drawings

The present invention is described referring now to the following drawings, wherein:

Fig. 1 shown use VNTI AlignX sequence multiple ratio to program (InforMax, Bethesda, the comparison of the S3a protein sequence that MD) carries out, wherein use the open point penalty in room be 10 and the room extend to 0.05 default setting.The residue of representing with white on the black background is identical; The residue of representing with white on the gray background is residue that guard or similar district, as the VNTI option is defined.

Fig. 2 has shown the sequence alignment from Lyamouri (Gene 294 (2002) 147-156) etc., it has shown from the high conservative in the S3a albumen of different plant species: homo sapiens (H.sapiens) (Hs), Rattus norvegicus (M.musculus) (Mm), Fugu rubripes (Temmincket Schlegel) (T.rubripes) (Tr), Africa xenopus (X.laevis) (Xl), drosophila melanogaster (D.melanogaster) (Dm), black fruit bat (D.virilis) (Dv), Caenorhabditis elegans (C.elegans) (Ce), yeast saccharomyces cerevisiae (S.cerevisiae) (Sc), Arabidopis thaliana (A.thaliana) (At), rice (O.sativa) is (Os) and H.holobium (Hh).Black shade is represented identical amino acid, and light gray is represented the amino-acid substitution guarded.The room is represented by dash.The conservative zone of topnotch is in the square frame.

Fig. 3 has shown binary vector, is used for expressing the S-phase specificity 40S S3a ribosomal protein gene (confidential reference items CDS0730) that the Arabidopis thaliana cyc07 be in (confidential reference items PRO0129) under the control of rice GOS2 promotor infers rice.

Fig. 4 has described the sequence that is used to carry out the inventive method in detail.

Embodiment

Invention will now be described by reference to the following examples, it only is for the purpose of illustration.

DNA operation: except as otherwise noted, recombinant DNA technology is according to (Sambrook (2001) Molecular Cloning:a laboratory manual, 3rd Edition Cold Spring HarborLaboratory Press, CSH, New York) or Ausubel grade in an imperial examination 1 volume and the 2nd roll up (1994), Current Protocols in Molecular Biology, the standard scheme of describing among the Current Protocols carries out.Be used for the standard material of plant molecular work and the Plant Molecular Biology Labfase (1993) that method is described in R.D.D Croy, publish by BIOS Scientific PublicationsLtd (UK) and Blackwell Scientific Publications (UK).

Embodiment 1: gene clone

Use Arabidopis thaliana seedling cDNA library (Invitrogen, Paisley, UK) as template, pcr amplification Arabidopis thaliana S3a (S-phase specificity 40S cyc07).After the RNA that extracts from seedling carried out reverse transcription, cDNA is cloned among the pCMV Sport 6.0.The average insertion size of gene pool is 1.5kb, and clone's original number is 1.59 * 10 ⁷Cfu.Original titre is defined as 9.6 * 10 ⁵Cfu/ml, and after first round amplification, become 6 * 10 ¹¹Cfu/ml.After plasmid extracted, the template of 200ng was used for 50 μ l PCR mixtures.Following primer is used for pcr amplification: primer prm02255 (justice, initiator codon is a black matrix, the AttB1 site is an italic: 5 ' GGGGACAAGTTTGTACAAAAAAGCAGGCTTCACAATGGCTGTCGGGAAGAA 3 ') and prm02256 (oppositely, complementary, terminator codon is a black matrix, the AttB2 site is an italic: 5 ' GGGGACCACTTTGTACAAGAAAGCTGGGTCCTAAGCTCCGATGATTTCT 3 '), it comprises the AttB site that is used for the Gateway reorganization.Use Hifi Tag archaeal dna polymerase, under standard conditions, carry out PCR.The PCR fragment of 789bp that increases, and use standard method to carry out purifying equally.Carry out the first step of Gateway method then, i.e. BP reaction, during this period, the PCR fragment is recombinated with the pDONR201 plasmid in vivo, according to the Gateway term, to produce " entry clone " p2782.Plasmid pDONR201 is available from Invitrogen, as the part of Gateway  technology.

Embodiment 2: vector construction

Entry clone p2782 then carries out the LR reaction with p0640, and p0640 is used for the purpose carrier that rice transforms.This carrier comprise be positioned at the T-DNA border with the lower section as functional element: plant selectable marker; The foliage filter screening mark; Be intended to and be cloned into aim sequence among the entry clone and carry out the Gateway expression cassette of the LR of reorganization in the body.The rice GOS2 promotor (PRO0129) that is used for constitutive expression is positioned at the upstream of this Gateway expression cassette.After the LR reconstitution steps, the expression vector as shown in Figure 3 that obtains is transformed in the soil Agrobacterium, and it is transformed in the rice plants subsequently.Make the rice plants growth of conversion, check the parameter described in the embodiment 3 then.

Embodiment 3: assessment and result

About 15 to 20 T0 rice transformants have independently been produced.Generation transformant is transferred to the greenhouse from tissue culture room and is grown, and results T1 seed.6 incidents are kept, and wherein separation in 3: 1 of transgenosis existence/shortage take place the T1 offspring.For in these incidents each,, selected about 10 to comprise genetically modified T1 seedling (abnormal shape and homozygote) and about 10 and lack genetically modified T1 seedling (invalid zygote) by the expression of monitoring visual indicia.Some T1 incident is used in generation with T1 at T2 and is further assessed for identical evaluation method.

Statistical analysis: F-test

The ANOVA of two factors (Mutability analysis) is used for the comprehensive evaluation of plant phenotype feature as statistical model.All measuring parameters with all plant of all incidents of gene transformation of the present invention are carried out the F test.Carry out the effect of F test with all transformation events of inspection gene pairs, and the total effect of check gene, also be referred to as whole genetic effect.The significance threshold value of true whole genetic effect is set to 5% probability level of F test.Significant F test value shows genetic effect, means to be not only the existence of gene or the difference that the position causes phenotype.

3.1 the parameter measurement that seed is relevant

Gather in the crops sophisticated elementary fringe, the bar code mark is carried out in pack, then at baking oven in 37 ℃ of dryings 3 days.To the fringe threshing, collect all seeds and counting then.Use the air-blowing device, full husk is separated from empty husk.Abandon the sky husk, and the part that keeps is counted once more.Full husk is weighed on analytical balance.This method produces seed correlation parameter group as described below.

Below table as a result shown the p value of the F test that is used for T1 assessment, T2 assessment, and the F of T1 that is used to unite and the T2 assessment p value of testting.When identical incident having been carried out two experiments, can consider Conjoint Analysis.This can be used for checking the consistence to the effect of two experiments, and increases the confidence level of conclusion.Used method is the mixture model method, and it considers the multilevel hierarchy (also i.e. experiment-incident-segregant) of data.The P value is by relatively likelihood ratio test and the distribution acquisition of card side.% difference between the transgenosis that each table all provides per generation and the corresponding invalid zygote.

3.1.1 full seed number

Number by the full husk of reservation after the counting separating step is determined full seed number.As shown in table 1 below, the p value of the F test of the T1 that is used to unite and T2 assessment is (the p value is 0.0071) significantly, and this existence that shows construct in the plant has remarkably influenced to the full seed number of transgenic plant.

Table 1

Full seed number
				% difference	The P value
T1	22	0.354
			T2	15	0.1829
Associating		0.0071

3.1.2 the seed overall yield of each plant

By to weighing, measure the seed overall yield from the whole full husk of plant results.As shown in table 2 below, the p value of the F test that the T1 that is used to unite and T2 estimate is (the p value is 0.0016) significantly, and this existence that shows construct in the plant has remarkably influenced to the seed gross weight of transgenic plant.

Table 2

The seed gross weight
				% difference	The P value
T1	29	0.0266
			T2	20	0.1037
Associating		0.0016

3.1.3 the harvest index of plant

Harvest index among the present invention is defined as seed overall yield and ground area (mm ²) between ratio, multiply by coefficient 10 again ⁶As shown in table 3 below, the p value of the F test that (and dividing other) T1 that is used to unite and T2 estimate is (the p value is 0.0005) significantly, and this existence that shows construct in the plant has remarkably influenced to the harvest index of transgenic plant.

Table 3

Harvest index
				% difference	The P value
T1	15	0.0358
			T2	14	0.0365
Associating		0.0005

3.1.4 thousand seeds heavy (TKW)

Infer this parameter from the full seed number and their gross weight of counting.As shown in table 4 below, the p value of the F test of the T1 that is used to unite and T2 assessment is (the p value is 0.0405) significantly, and this existence that shows construct in the plant has remarkably influenced to the TKW of transgenic plant.

Table 4

Thousand seeds are heavy
				% difference	The P value
T1	3	0.3353
			T2	2	0.1257
Associating		0.0405

Sequence table

＜110〉Cropdesign NV

＜120〉have the plant of growth characteristics of improvement and the method for preparing described plant

<130>CD-112-PCT

<150>EP 04101179.2

<151>2004-03-22

<150>US 60/556,847

<151>2004-03-28

<160>6

<170>PatentIn version 3.3

<210>1

<211>830

<212>DNA

＜213〉Arabidopis thaliana (Arabidopsis thaliana)

<400>1

atggctgtcg ggaagaacaa gaggatttca aagggtagga aaggaggaaa gaagaaggct 60

gttgatccct tctccaagaa ggattggtat gacgtgaagg ctcctggttc tttcacgaac 120

aggaatgttg ggaagactct tgtttccagg actcagggta ccaagattgc ctctgaggga 180

ctgaaacaca gggtgtttga ggtttctctt gctgatctac aaaatgatga ggataatgcc 240

tacaggaaga tccgtcttag agctgaagat gttcagggaa ggaatgtgtt gacccagttc 300

tggggtatgg atttcacaac cgacaagcta aggtcattgg tgaagaagtg gcagactttg 360

attgaagccc atgtcgatgt gaaaaccaca gacggctaca ccttgaggat gttctgcatc 420

gccttcacaa agagacgtgc taaccaagtg aagcgtacct gttacgctca atccagccaa 480

atccgtcaga tccgcagaaa gatgagtgag attatggtga aggaggcttc atcttgtgac 540

ctcaaggagc tagtggccaa gttcatccca gaggccattg gaagagagat tgagaaggca 600

acacagggca tctacccgtt gcagaatgtg ttcatccgta aagtgaagat cctaaaggct 660

cccaagtttg accttggaaa gctcatggag gtgcatggag attacacagc agaggatgtt 720

ggtgtgaagg tagacaggcc agctgatgag acaatggttg aggagccaac agaaatcatc 780

ggagcttagg ggattataga tttgtttgtt ttttcgctgg caaaaaaaaa 830

<210>2

<211>261

<212>PRT

＜213〉Arabidopis thaliana

<400>2

Met Ala Val Gly Lys Asn Lys Arg Ile Ser Lys Gly Arg Lys Gly Gly

1 5 10 15

Lys Lys Lys Ala Val Asp Pro Phe Ser Lys Lys Asp Trp Tyr Asp Val

20 25 30

Lys Ala Pro Gly Ser Phe Thr Asn Arg Asn Val Gly Lys Thr Leu Val

35 40 45

Ser Arg Thr Gln Gly Thr Lys Ile Ala Ser Glu Gly Leu Lys His Arg

50 55 60

Val Phe Glu Val Ser Leu Ala Asp Leu Gln Asn Asp Glu Asp Asn Ala

65 70 75 80

Tyr Arg Lys Ile Arg Leu Arg Ala Glu Asp Val Gln Gly Arg Asn Val

85 90 95

Leu Thr Gln Phe Trp Gly Met Asp Phe Thr Thr Asp Lys Leu Arg Ser

100 105 110

Leu Val Lys Lys Trp Gln Thr Leu Ile Glu Ala His Val Asp Val Lys

115 120 125

Thr Thr Asp Gly Tyr Thr Leu Arg Met Phe Cys Ile Ala Phe Thr Lys

130 135 140

Arg Arg Ala Asn Gln Val Lys Arg Thr Cys Tyr Ala Gln Ser Ser Gln

145 150 155 160

Ile Arg Gln Ile Arg Arg Lys Met Ser Glu Ile Met Val Lys Glu Ala

165 170 175

Ser Ser Cys Asp Leu Lys Glu Leu Val Ala Lys Phe Ile Pro Glu Ala

180 185 190

Ile Gly Arg Glu Ile Glu Lys Ala Thr Gln Gly Ile Tyr Pro Leu Gln

195 200 205

Asn Val Phe Ile Arg Lys Val Lys Ile Leu Lys Ala Pro Lys Phe Asp

210 215 220

Leu Gly Lys Leu Met Glu Val His Gly Asp Tyr Thr Ala Glu Asp Val

225 230 235 240

Gly Val Lys Val Asp Arg Pro Ala Asp Glu Thr Met Val Glu Glu Pro

245 250 255

Thr Glu Ile Ile Gly

260

<210>3

<211>1169

<212>DNA

＜213〉sugarcane (Saccharum officinarum)

<400>3

aagatgaagc ctttgtcatg gtgcatgcta aagatgctga ggctgagaag ttgagggatg 60

aaccatgaca aaggcttcat ctttctcgac ctgaatcctg tccacattcc ccttcagcat 120

cttcaattca gcctcgatca ttttcttctt aagcaccccg ccgtcgttct cttcctgcat 180

ccccgcccca ttccctagcg tcgcccccct cgccgccgca cggacgcagc gacgagctct 240

cgcagcagca atggcggttg gcaagaataa gcgtatctcc aagggcaaga agggaggcaa 300

gaagaagacc gtggatccgt tcagcaaaaa ggattggtat gatatcaagg ctccgtcggt 360

cttcagcgtg cgcaacatcg gcaagaccct ggtctccagg acacagggca ccaagattgc 420

ctctgagggt ttaaagcaca gagtatttga ggtctccttg gctgatcttc agagtgatga 480

agaccaggcg tacaggaaga tcagacttcg tgcagaggat gtacaaggga gaaatgttct 540

cacaaacttc tggggtatga gcttcaccac cgacaagctc cgttcacttg tgaagaagtg 600

gcagacgctt attgaggctc atgttgatgt caagaccacc gataactata tgctgcggct 660

gttctgcatt gggttcacca agaggcggcc caatcaagtg aagcgcactt gctatgctca 720

agcaagccaa atcagacaga ttcgtcggaa gatgactgaa atcatgagca accaagcttc 780

aacttgtgat ctgaaagagc tcgtgtccaa gttcatccct gaggtcattg gaaaggaaat 840

cgagaaagcc acctctagca tattcccctt gcaaaatgtc ttcatccgca aggtgaagat 900

cctgaaagca ccaaagttcg acattggaaa gctcatggag gtccatggtg actatgccaa 960

ggaggatgtt ggtgtcaaga tggacaggcc tgctgaaggc gacgaggcca tgggaggaca 1020

ggaggttgct gcagctgagt gattagtctc actgtttacg tccgagttag agctgccata 1080

tttccttgaa acacttagga acactttttt tgagagtctg acatgtggtg gcttcgattc 1140

tccttgaaaa tttgcagcat gggaaatgt 1169

<210>4

<211>263

<212>PRT

＜213〉sugarcane

<400>4

Met Ala Val Gly Lys Asn Lys Arg Ile Ser Lys Gly Lys Lys Gly Gly

1 5 10 15

Lys Lys Lys Thr Val Asp Pro Phe Ser Lys Lys Asp Trp Tyr Asp Ile

20 25 30

Lys Ala Pro Ser Val Phe Ser Val Arg Asn Ile Gly Lys Thr Leu Val

35 40 45

Ser Arg Thr Gln Gly Thr Lys Ile Ala Ser Glu Gly Leu Lys His Arg

50 55 60

Val Phe Glu Val Ser Leu Ala Asp Leu Gln Ser Asp Glu Asp Gln Ala

65 70 75 80

Tyr Arg Lys Ile Arg Leu Arg Ala Glu Asp Val Gln Gly Arg Asn Val

85 90 95

Leu Thr Asn Phe Trp Gly Met Ser Phe Thr Thr Asp Lys Leu Arg Ser

100 105 110

Leu Val Lys Lys Trp Gln Thr Leu Ile Glu Ala His Val Asp Val Lys

115 120 125

Thr Thr Asp Asn Tyr Met Leu Arg Leu Phe Cys Ile Gly Phe Thr Lys

130 135 140

Arg Arg Pro Asn Gln Val Lys Arg Thr Cys Tyr Ala Gln Ala Ser Gln

145 150 155 160

Ile Arg Gln Ile Arg Arg Lys Met Thr Glu Ile Met Ser Asn Gln Ala

165 170 175

Ser Thr Cys Asp Leu Lys Glu Leu Val Ser Lys Phe Ile Pro Glu Val

180 185 190

Ile Gly Lys Glu Ile Glu Lys Ala Thr Ser Ser Ile Phe Pro Leu Gln

195 200 205

Asn Val Phe Ile Arg Lys Val Lys Ile Leu Lys Ala Pro Lys Phe Asp

210 215 220

Ile Gly Lys Leu Met Glu Val His Gly Asp Tyr Ala Lys Glu Asp Val

225 230 235 240

Gly Val Lys Met Asp Arg Pro Ala Glu Gly Asp Glu Ala Met Gly Gly

245 250 255

Gln Glu Val Ala Ala Ala Glu

260

<210>5

<211>51

<212>DNA

＜213〉artificial sequence

<220>

＜223〉primer prm02255

<400>5

ggggacaagt ttgtacaaaa aagcaggctt cacaatggct gtcgggaaga a 51

<210>6

<211>49

<212>DNA

＜213〉artificial sequence

<220>

＜223〉primer prm02256

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ggggaccact ttgtacaaga aagctgggtc ctaagctccg atgatttct 49

Claims (24)

1. a method that improves plant growth characteristics is included in the nucleic acid of introducing in the plant and expressing coding small subunit ribosomal protein S3a polypeptide.

2. the method for claim 1 is included in and introduces in the plant and express:

(i) part of S3a coding nucleic acid;

(ii) can with the sequence of S3a coding nucleic acid hybridization;

The (iii) splice variant selected of S3a coding nucleic acid;

The (iv) allelic variant of S3a coding nucleic acid; With

(v) the encode homologue of S3a aminoacid sequence or the nucleic acid of derivative,

Wherein, (i) to (v) coded protein and known S3a protein sequence are compared and comprised and the corresponding conservative region in frame district shown in Fig. 2 sequence alignment.

3. claim 1 or 2 method, the plant growth characteristics of wherein said improvement is the productive rate of increase for the corresponding wild-type plant.

4. the method for claim 3, the productive rate of wherein said increase are the seed productive rates that increases.

5. claim 3 or 4 method, the productive rate of wherein said increase is selected from the seed biomass that (i) increases; (ii) (full) seed number of Zeng Jiaing; The (iii) seed size of Zeng Jiaing; The (iv) seed volume of Zeng Jiaing; (the v) harvest index of Zeng Jiaing; (vi) thousand seeds that increase weigh (TKW).

6. each method in the claim 1 to 5, the plant growth characteristics of wherein said improvement are the growth velocitys that increases.

7. each method in the claim 1 to 6, the nucleic acid of wherein said coding S3a derives from plant, preferably derives from dicotyledons, also preferably derives from Cruciferae, and more preferably this nucleic acid comes from Arabidopis thaliana.

8. each method in the claim 1 to 7, the nucleotide sequence of wherein said coding S3a are crossed in plant and are expressed.

9. each method in the claim 1 to 8, the expression of nucleic acids of wherein said coding S3a is by constitutive promoter especially GOS2 promoters driven.

10. be used to improve the especially productive rate method of seed productive rate particularly of plant growth characteristics, be included in the genetic modification of the locus of the gene of introducing coding S3a polypeptide or its variant in the plant.

11. the method for claim 10, wherein said genetic modification is realized by one of following: site-directed mutagenesis, homologous recombination, TILLING, directly evolve and T-DNA activates.

12. can be by the plant of each method acquisition in the claim 1 to 11.

13. construct, it comprises:

(i) S3a coding nucleic acid;

(ii) can drive one or more control sequences that (i) amplifying nucleic acid sequence is expressed; Randomly

(iii) transcription termination sequence.

14. the construct of claim 13, wherein said control sequence comprises at least one constitutive promoter, preferred GOS2 promotor.

15. construct plant transformed with claim 13 or 14.

16. be used to produce the method for the transgenic plant of the growth characteristics with improvement, this method comprises:

(i) the S3a coding nucleic acid is incorporated in plant or the vegetable cell;

(ii) promoting regeneration and making culturing plants cell under the sophisticated condition of plant-growth.

17. have the transgenic plant of the growth characteristics of improvement, be characterised in that and in described plant, introduce and express the proteic nucleotide sequence of coding S3a.

18. claim 12,15 or the transgenic plant of claim 17, wherein said plant is a monocotyledons, as sugarcane, or be selected from the cereal class of rice, corn, wheat, barley, soybean, Chinese sorghum, Sunflower Receptacle, rape, sugarcane, clover, grain, barley, Semen Brassicae campestris and cotton.

19. the part gathered in the crops of each plant in the claim 12,15,17 or 18.

20. the part gathered in the crops of the plant of claim 19, wherein said gather in the crops the part be seed.

21.S3a or the S3a coding nucleic acid especially improves the particularly purposes in the seed productive rate of productive rate in the growth characteristics of improvement plant.

22. the purposes of claim 21, wherein said seed productive rate comprise as the next item down or multinomial: (full) seed number of increase, the seed weight of increase, the harvest index of increase and the TKW of increase.

23. the purposes of claim 21 or 22, wherein said S3a coding nucleic acid is shown in SEQ ID NO:1, and wherein said S3a is the amino acid shown in SEQ ID NO:2.

24.S3a or the S3a coding nucleic acid is as the purposes of molecular marker.

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