CN107893082A - Rice leaf sugar accumulation related gene SAC1 and its SAC1 mutator and application - Google Patents

Rice leaf sugar accumulation related gene SAC1 and its SAC1 mutator and application Download PDF

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CN107893082A
CN107893082A CN201711463928.3A CN201711463928A CN107893082A CN 107893082 A CN107893082 A CN 107893082A CN 201711463928 A CN201711463928 A CN 201711463928A CN 107893082 A CN107893082 A CN 107893082A
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sac1
leu
ser
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ala
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何光华
朱小燕
桑贤春
李云峰
王楠
赵芳明
张长伟
凌英华
杨正林
吴仁鸿
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Southwest University
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Abstract

The present invention relates to a kind of rice leaf sugar accumulation related gene SAC1 and its SAC1 mutator and application,Wherein rice leaf sugar accumulation related gene SAC1 nucleotide sequence is as shown in SEQ ID No.13,Amino acid sequence is as shown in SEQ ID No.14,And SAC1 mutators are converted to A in the bit base of encoder block the 1238th compared with wild type by G,The coded amino acid of the 413rd is caused by arginine (Arg) variation to be lysine (Lys),Yellow green and aging in advance is presented on time of infertility blade top in rice SAC1 mutant after the gene mutation,And plant is downgraded,Genetic analysis finds that the character is dominant character,Field removal of impurities and Purity Identification can be carried out using this character as molecular labeling,It is significant to the genetic breeding of rice.

Description

Rice leaf sugar accumulation related gene SAC1 and its SAC1 mutator and application
Technical field
The invention belongs to genetic arts, are related to rice leaf sugar accumulation related gene SAC1, further relate to Rice Leaf piece candy Accumulate mutator and the application of related gene.
Background technology
Rice (Orvza sativa L.) is most important cereal crops in the world, is planted extensively 120 countries and regions Plant, the global population of more than half is using rice as staple food.China's success improved crossing rice, and promotion and application are carried out, increase production Grain realize that foodgrain self-sufficiency creates condition for some countries, outstanding contribution has been made safely to country or even world food. Hybrid seed caused by yet with sterile line fertility instability mixes, and has a strong impact on the life of hybrid purity and hybrid rice Production, or even to heavy losses caused by peasant, restrict hybrid paddy rice and play a greater role.In two systems and ternary hybrid rice breeding and life In production practice, it has been found that sterile line sterility unstable expression phenomenon generally existing.Therefore, how parent propagation is effectively improved Dust removal rate during the production of hybrid seeds, quick and precisely identify in seed production process because the seed purity that fertility fluctuation is likely to occur is asked Topic a, it has also become important topic in hybrid rice research and application.Leaf color mutant can be used as seedling stage mark property to carry out Crop hybrid generation breeding of new variety and Purity Identification, its advantage is can be by observing the presence of mark property in seedling stage Or whether disappear to identify true and false hybrid, therefore it is pure in three systems and double-linear hybrid rice parent propagation, hybrid seeding, hybrid seed Degree identification etc. is fully used.This technology is intuitively accurate, easy quick, have the identification of in general Planting in the different location and The incomparable superiority of DNA molecular marker identification technology.The man of breeding in recent years has selected multiple with leaf colour marker Sterile line, as albefaction turns greenery color marker sterile line rich A, yellowish leaf sterile line mark 810S, yellow leaf sterile line topaz A and yellowish green in vain The yellow mark A series of leaf sterile line.But because most seedling stage mark property monosystem characters itself are not excellent, photosynthetic efficiency declines, cause Vine growth and development is abnormal and the underproduction, is restricted its application in the practices of breeding.Thus, find some stable something lost Biography, leaf variegation are extremely important to other characters Leaf color mutant that especially yield, quality trait have no significant effect.
The content of the invention
In view of this, an object of the present invention is to provide a kind of rice leaf sugar accumulation related gene SAC1, this hair The bright second purpose is to provide rice leaf sugar accumulation gene SAC1 mutators, and the gene is time of infertility mark property, Strong instrument is provided for Transgenic Rice research, promotes hybrid rice breeding research；The third object of the present invention is to provide Application of the SAC1 mutators in the molecular breeding of rice leaf sugar accumulation character.
To achieve the above object, the present invention utilizes extensive No. 10 of ethylmethane sulfonate (EMS) mutagenesis Elite restorer line red silk, obtains One time of infertility blade top shows as yellow green and accumulates the mutant (being named as sac1) of a large amount of sugar, genetic analysis It was found that the mutant character is controlled by a pair of dominant karyogenes (being named as SAC1).Molecular mapping result shows that the gene is determined Between the 7th the short arm of a chromosome end SSR marker c7sr2 and Indel Tag ID 10 in 7.48kb section.
On the basis of said gene positioning, the present invention is predicted by candidate gene, Homology search and gene order difference Compare, it is expressing protein (Loc_Os07g02520) to have primarily determined that SAC1；Then, the present invention is with extensive No. 10 of rice wild type red silk It is material with rice leaf sugar accumulation mutant sac1, has cloned rice leaf sugar accumulation related gene SAC1 and SAC1 mutation base Cause, rice leaf sugar accumulation related gene SAC1 have the nucleotide sequence as shown in SEQ ID No.13, as a result shown, SAC1 Gene is free of introne, is only made up of 1 extron, and CDS encoder blocks total length is 2361bp, encodes 787 amino acid, its ammonia altogether Base acid sequence is as shown in SEQ ID No.14.The nucleotide sequence of SAC1 mutators is extensive with red silk as shown in SEQ ID No.23 No. 10 wild type genes are compared, and SAC1 mutators are converted to A in the bit base of encoder block the 1238th by G, cause the 413rd coding Amino acid by arginine (Arg) variation be lysine (Lys), the amino acid of acquisition is as shown in SEQ ID No.24.
Then, the present invention constructs overexpression vector and rice transformation blade sugar accumulation mutant sac1, character analysis hair Existing transgenic positive plant height recovers normal, and normal green is recovered on blade top, and expression analysis shows transgenic positive plant Middle SAC1 expression quantity also significantly improves.Accordingly, it is determined that mutant character is as caused by SAC1 gene mutations.
Therefore, application of the SAC1 mutators in the molecular breeding of rice leaf sugar accumulation character.Preferably, the water The kind of rice is extensive No. 10 of red silk.
The beneficial effects of the present invention are：The invention provides rice leaf sugar accumulation related gene SAC1 and SAC1 mutation Gene, the gene are seedling stage mark property, and the Other Main Agronomic Characters such as setting percentage, mass of 1000 kernel are generated and significantly affected, and are rice Genetic breeding research provides strong instrument, is laid the foundation for the purebred sterile line of seed selection.
Brief description of the drawings
In order that the purpose of the present invention, technical scheme and beneficial effect are clearer, the present invention provides drawings described below and carried out Explanation：
Fig. 1 is extensive No. 10 of wild type red silk and rice leaf sugar accumulation mutant sac1 morphological observations (A-B：Tiller Extensive No. 10 of phase phase wild type red silk and sac1 mutant plants and blade；C：Extensive No. 10 of maturity period wild type red silk is planted with sac1 mutant Strain；D：Extensive No. 10 of maturity period wild type red silk is mutated body segment and fringe with sac1).
Fig. 2 be extensive No. 10 of wild type red silk and rice leaf sugar accumulation mutant sac1 plant and blade from 2 leaf phases to 6 leaf phases The starch coloration result (1 of morning and evening：2 leaves early stage；2：2 leaf late periods；3：3 leaf late periods；4：4 leaf late periods；5：5 leaf late periods；6：6 leaves evening Phase).
Fig. 3 contains for the analysis of extensive No. 10 of wild type red silk and rice leaf sugar accumulation mutant sac1 cell ultrastructure, sugar Measure fixed (A-B：The cell ultrastructure analysis that wild type red silk is extensive No. 10；C-D：The cell ultrastructure analysis of sac1 mutant； E-K：The content of extensive No. 10 of wild type red silk and rice leaf sugar accumulation mutant sac1 leaf starch, sucrose and glucose).
Fig. 4 is that extensive No. 10 of wild type red silk analyzes (A with sac1 mutant Other Main Agronomic Characters：Plant height；B：Spike length；C：It is solid Rate；D：Mass of 1000 kernel；E：Primary branch number and Secondary branch number).
Fig. 5 is heredity and the physical map of rice leaf sugar accumulation mutator SAC1 genes, and wherein A is the preliminary of SAC1 Positioning, between the 7th chromosome breaks arm SSR marker c7sr1 and InDel Tag ID -10；B is the finely positioning of SAC1 genes, 7.48kb scope and mutant sac1 candidate gene Loc_ between SSR marker c7sr2 and InDel Tag ID -10 Os07g02520 structure and mutated site.
Fig. 6 is that the overexpression phenotypic evaluation of sac1 mutant is analyzed, and wherein A is wild type WT, mutant sac1 and super table Up to transgenic positive plant (OESAC1-C) phenotypic analysis；B is SAC1 genes in wild type WT, mutant sac1 and overexpression Expression analysis in transgenic positive plant (OESAC1-C).
Embodiment
Below in conjunction with accompanying drawing, the preferred embodiments of the present invention are described in detail.
The material used in the embodiment of the present invention：Wild rice material red silk extensive No. 10 (WT) and rice leaf sugar accumulation are dashed forward Variant sac1 is cultivated by Southwest University's rice research；M-MLV reverse transcriptases, high-fidelity DNA polymerase PFU, Taq DNA gather Synthase, T4DNA ligases, restriction enzyme, pMD19-T carriers, Trizol kits, DNA gel QIAquick Gel Extraction Kit, plasmid Extracts kit, λ-Hind III DNA Marker and DL5 000DNA Marker are purchased from TaKaRa companies；DNA Marker III is purchased from TIANGEN Biotech (Beijing) Co., Ltd.；Ampicillin (Ampicillin, Amp) and kalamycin (Kanamycin, Kan) is Sigma Products；The measure kit of carbohydrate and starch is purchased from Hayward companies；Primer synthesizes Completed with DNA sequencing by the handsome Bioisystech Co., Ltd in Shanghai；Other chemical reagent are limited purchased from Beijing ancient cooking vessel state biotechnology Responsible company；Bacillus coli DH 5 alpha, Agrobacterium LBA4404 are preserved by this laboratory.
Embodiment 1, rice leaf sugar accumulation mutant sac1 acquisition and morphological observation
It is found that a time of infertility blade top is shown as in rice research institute of Southwest University rice EMS mutant library Yellowish green leaf and with the mutant of early ageing, sac1 (Fig. 1) is named as according to phenotypic characteristic.The character is observed through excessively generation, table Now stablize heredity, extensive No. 10 of transmission electron microscope observing seedling stage wild type red silk and sac1 mutant blade cell ultra microstructures, mutant Eucaryotic cell structure is complete, and amylum body increases, and chloroplast grana lamellar structure is by destruction (A-F in Fig. 3)；Other Main Agronomic Characters such as fringe Length, number of grain per ear, setting percentage, Primary branch number, Secondary branch number and mass of 1000 kernel etc. substantially reduce, and have had a strong impact on the battalion of rice Health length and reproductive growth (Fig. 4).
Embodiment 2, leaf starch dyeing and sugared content measure
Until 6 leaf phases since 2 leaf phases, the blade of extensive No. 10 of wild type red silk and sac1 mutant is taken respectively, using iodo- iodine Change potassium decoration method to dye it, extensive No. 10 of 6 wild type red silks and sac1 are by the unnecessary TP of the photosynthesis of one day at night Synthetic starch is stored in blade, is dyed to blueness；6 points of morning, the leaf starch that wild type red silk is extensive No. 10 pass through a night almost Decompose completely, and then the starch of Different sites of leaf decomposes situation difference in sac1, blade dyes blueness (figure by different degrees of 2).No. 10 extensive to wild type red silk and sac1 starch and soluble sugar content measure shows, starch, sucrose and the Portugal of sac1 mutant Grape sugared content is respectively 940%, 120% and 130% (E-K in Fig. 3) of extensive No. 10 of wild type red silk.
Embodiment 3, the analysis of mutation sac1 gene genetics and positioning
Hybridized with the western agriculture 1A (Xinong1A) of the normal sterile line of phenotype and sac1 mutant, all F1Plant occurs certain Trait segregation.F2There is obvious parents' character segregation phenomenon in colony, in the 14315 yellowish green and early ageing in group of hill body middle period top 10812 plants of individual plant, 3503 plants of normal individual plant, through card square test, meet 3:1 segregation ratio, show the mutant character by a pair Dominant gene controls.
Primary Location：Choose the 400 pairs of SSR markers amplification parent being uniformly distributed on 12 chromosomes of rice and gene pool DNA, as a result find that the mark RM20776 and RM6663 that are located at the 7th chromosome may be chain with SAC1, its genetic distance is respectively 7.6cM and 4.6cM (A in Fig. 5).
Finely positioning：According to the rice variety 93-11 sequences announced, enter one between RM20776 and RM6663 is marked Step screening and develop 10 couples of In/Del mark and 7 pairs of SSR markers, wherein ID-9, ID-10, RM20848, c7sr1, c7sr2 and RM6222 shows polymorphism (table 1) between parent.All F are further analyzed with this 6 pairs of primers2Recessive mutation individual plant, finally Between SAC1 is positioned at into SSR marker c7sr2 and Indel Tag ID -10, physical distance be 7.48kb in the range of (in Fig. 5 B)。
Table 1, the InDel flags sequence with polymorphism
Only 1 ORFs (http between In/Del Tag IDs -3 and SSR marker RM6222:// Www.gramene.org), by Homology search and gene order comparative analysis, it is found that function does not annotate gene in the section (Loc_Os07g02520), it may be target gene.The gene does not include introne, is only made up of an extron, CDS codings Frame total length is 2361bp, and its nucleotide sequence encodes 787 amino acid, amino acid sequence is such as altogether as shown in SEQ ID No.13 Shown in SEQ ID No.14.
Embodiment 3, clone's Loc_Os07g02520 genes
It is soft using Vector NTI according to the listed rice Nipponbare gene Loc_Os07g02520 sequences of GenBank The mRNA special primers of extensive No. 10 of part design amplification wild type red silk and sac1 mutant Loc_Os07g02520 sequences：Sense primer SAC1-F：5’-atggagcaacatgtggaacg-3’(SEQ ID No.15)；Anti-sense primer SAC1-R：5’- tcacactgtcccggctgca-3’(SEQ ID No.16).Extensive No. 10 of wild type red silk and mutant sac1 is taken to be trained in illumination respectively The spire 2g of two weeks is supported, is put into grind into powder in liquid nitrogen rapidly, total serum IgE is extracted according to Trizol kit specifications.Gained The electrophoresis result of extensive No. 10 of wild type red silk and mutant sac1 total serum IgEs shows master tape complete display, 28S and 18S band brightness Than being about 2:1, the concentration and purity for illustrating RNA meet requirement of experiment, can be used for synthetic double chain cDNA.Then respectively with gained Extensive No. 10 of wild type red silk and mutant sac1 total serum IgEs are template, according to M-MLV reverse transcriptase specifications, are drawn using Oligo (dT) Thing carries out reverse transcription and obtains cDNA；Again using cDNA as template, using sequence shown in SEQ ID No.15 and SEQ ID No.16 as spy Different primer and high-fidelity DNA polymerase PFU enters performing PCR amplification, and PCR reaction conditions are：94 DEG C of pre-degenerations 5 minutes；Then 94 DEG C Denaturation 30 seconds, 55 DEG C of renaturation 30 seconds, 72 DEG C extend 2.5 minutes, totally 35 circulations；Last 72 DEG C extend 10 minutes.By RT-PCR Product carries out the detection of 1.0% (g/mL) agarose gel electrophoresis.As a result show, extensive No. 10 of wild type red silk and mutant sac1 are expanded Product is being in single specificity band about at 2000+bp, and extensive No. 10 amplified productions of wild type red silk are named as into SAC1 genes, Mutant sac1 amplified productions are named as SAC1 mutators (SAC1 ').
Then gel extraction purifying, the SAC1 genes and SAC1 of purifying are carried out according to DNA gel QIAquick Gel Extraction Kit specification Mutator is connected overnight with pMD 19-T carriers in the presence of T4DNA ligases in 16 DEG C, connection product conversion large intestine bar Bacterium DH5 α competent cells, with the LB plate screening positive colony containing ampicillin, plasmid is extracted, is sequenced after PCR identifications, Recombinant vector pMD 19-T-SAC1 and pMD 19-T-SAC1 ' are obtained respectively.By recombinant vector pMD 19-T-SAC1 and pMD19-T- SAC1 ' send sequencing company to be sequenced, and SAC1 mutant gene sequences is as a result shown as shown in SEQ ID No.23, with wild type red silk Extensive No. 10 are compared, and SAC1 genes are converted to A in the bit base of encoder block the 1238th by G, cause the coded amino acid of the 413rd by essence Propylhomoserin (Arg) variation be lysine (Lys), after mutation amino acid sequence as shown in SEQ ID No.24, SAC1 gene orders and Nipponbare gene Loc_Os07g02520 sequences are consistent.
The functional verification of embodiment 4, mutator SAC1
In order to verify that rice mutant sac1 blade sugar accumulation and leaf top yellow character are drawn by mutator SAC1 Rise, the extensive No. 10 SAC1 genes of wild type red silk, structure overexpression vector and rice transformation blade sugar are expanded using primer shown in table 2 Accumulation and leaf top chlorophyll-reduced mutant sac1, character analysis find that transgenic positive plant leaf recovers normal green, chlorophyll Content analysis shows that transgenic positive plant leaf Chlorophyll content recovers normal level (A in Fig. 6).Then utilize in table 3 Primer SEQ ID No.19 and SEQ ID No.20 to SAC1 genes in wild type and overexpression transgenic positive plant sac1 Quantitative fluorescence analysis is carried out, while is by internal reference reaction system of UBQ5：2 μ L cDNA moulds are added in 25 μ L reaction system Plate, 2 μ L primers, 12.5 μ L SYBR Green fluorescent dyes and 8.5 μ L RNase-free H2O, it is glimmering in ABI Prism 7500 Fluorescent quantitation amplification is carried out on Fluorescent Quantitative PCR instrument；Amplification condition is：94 DEG C of pre-degenerations 5 seconds；94 DEG C be denatured 5 seconds, 60 DEG C 30 seconds, 40 circulations；And solubilization solution curve 65 DEG C 5 seconds, 95 DEG C 0.5 DEG C, then utilize CFX-Manager softwares to carry out the collection of data With processing.As a result find that SAC1 gene expression amounts substantially raise (B in Fig. 6) in overexpression transgenic positive plant.Accordingly, it is determined that Mutant blade sugar accumulation and leaf top yellow character are as caused by SAC1 gene mutations.
Table 2, for building overexpression vector primer and restriction enzyme site
Table 3, the primer sequence for qRT-PCR
Gene Positive sequence (5 ' → 3 ') Reverse sequence (5 ' → 3 ')
Q-SAC1 gggctgatgcagtcatcgctgg(SEQ ID No.19) ctccgcagcctggtcgggag(SEQ ID No.20)
UBQ5 accacttcgaccgccactact(SEQ ID No.21) acgcctaagcctgctggtt(SEQ ID No22)
The bioinformatic analysis of embodiment 6, rice leaf sugar accumulation and leaf top yellow aging mutator SAC1
SAC1 gene orders are utilized into the ORF Finder (http in NCBI://www.ncbi.nlm.nih.gov/ Gorf/gorf.html ORFs identification) is carried out.As a result show, SAC1 genes are by a complete and continuous open reading Frame forms.
SAC1 gene coded protein structures are predicted and analyzed with SMART softwares, show SAC1 gene coded proteins N- contains membrane spaning domain in end, and the albumen includes two conservative unknown function domain DUF4220 and DUF594, and this two The synchronous appearance frequent in plant of individual domain.
Leaf color mutant excellent using Comprehensive Traits, and being controlled by dominant karyogene, will by the means such as hybridizing, being returned In the sterile line of the system of Leaf color mutant channel genes three or two systems, the removal of impurities pure keeping efficiency in parent's reproductive process can be effectively improved； Also the seed purity problem being likely to occur in seed production process by fertility fluctuation can quick and precisely be identified.So the present invention discloses SAC1 mutators important genetic resources is provided for the molecular breeding of rice.Plant table is built based on SAC1 mutators Up to carrier and the rice sterile line of high-quality background is converted, then the rice cell of conversion is cultivated into yellowish green leaf sterile line, you can be logical Cross transgenosis and quickly realize yellowish green leaf sterile line.Seedling stage, yellowish green leaf mark property can be by observing depositing for mark property in seedling stage Or whether disappear to identify true and false hybrid, therefore in three systems and double-linear hybrid rice parent propagation, hybrid seeding, hybrid seed Purity etc. is fully used.This technology has the characteristics of quick, simple, accurate, inexpensive, hence it is evident that better than one As Planting in the different location identification and DNA molecular marker identification technology.
Finally illustrate, preferred embodiment above is merely illustrative of the technical solution of the present invention and unrestricted, although logical Cross above preferred embodiment the present invention is described in detail, it is to be understood by those skilled in the art that can be Various changes are made to it in form and in details, without departing from claims of the present invention limited range.
Sequence table
<110>Southwest University
<120>Rice leaf sugar accumulation related gene SAC1 and its SAC1 mutator and application
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gcctctttga atcgcaggat t 21
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gtgtgtgtgc tccaacaaac tg 22
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ttctacgccg tcaccttcgg atcctgccgc cgctggtcta gccgatggat catccaaaag 180
gttttcctgg ctgcgaatgc gctgtctgtg tccctaggaa catacagcat agggctgatg 240
cagtcatcgc tggtgaagag caagatgtac cctgtgtggt cggtgtcatt gcttgccctc 300
tttgctagca ttgactccat caacacattc ggcctcgact atagtggcca attcctgaag 360
ctgttgtatc agctcttcct ctactttggg tatgtgcttg tgatcagcat ctccggcaat 420
accaataatg gcatggtggc cgctgcgatt ggcatgcttt gcagcgtcac cctatgtaag 480
ggattccata ggctgatggc ctatgtgctc ccgaccaggc tgcggagcat gaacaaagag 540
attgcagatc gtatggaagt ggtgaagggg gggccaaact ttaacgcggc catcatgcaa 600
gggtatcaat acctggttga taagacttat tatccatcca taaccctgga agaattgtgg 660
aagcctgaca atggcatgga caaactagga acagatgctg atgcctacaa ggacatatgc 720
ctctctttct ccctctgtca cctgctacag cgacgcttct ttggttttag ttgtgctgag 780
tctgaccggc cagagacccg tagctttgtg ttggaaggct tgcttcaacc tgtagcttca 840
agtagttcat catcacaagg tactggtggt tctcgtggta actacgagag ggccttcaag 900
gtgattgagg ttgagctagc cttcatgtac gactacatct acagcagcaa cgccttcgtc 960
cattactatg aagctggagc ttgcaccgcc tgggcaatag cttcaatcct gaccacatgt 1020
tttctatgtg tggctgttgc cctacagcaa aaagaagcca ttattgcatc atctgcggtc 1080
actatgttgg actcggaggt aattagcaca acgatgacac cagacattgt catcaccctt 1140
gtcatactgg tgtcccttgc ttccctccaa gtgttgcagc tggtgaactc cttgtcgtcc 1200
aattgggcaa gagtctccct tgtctgtcat cttttcagga gaaatgccaa caacctggag 1260
ccatcgatgg gcttgaagct gaggttgttc ctttccagga tcaaactatt ggacaaatac 1320
cagtggcaga acaagcttgg gcagtacgag agctcatcgt ggctcaagaa agctctaggg 1380
tttgtcagct tcaagatagg acaatgttgc tataagccag tgtttcattt catgctctac 1440
tgcgtgcggt gcatgatgcg atggccaacc atttgctgcc catgctatgt ggtgtgctct 1500
ccgctctgca actgggtgtt caaattgcag cctagttggc agttgctccc aggcccttgt 1560
tgggtgatat actccgccct ctacaagttg tttggcctgc agtacattca acaggtgctg 1620
acagacatgt tgggcattag cactgcaagc tccattcagc tgcccattga ggtgaagagt 1680
accgtgattg atgccctcgt tgggatcctc atgcccaggc ccaaccatga cagtgtcgtg 1740
ttgtccagtg gatcaacatc gctagccaag aatggcttgc aggacaagtt cataaggcag 1800
tacgcttcga caagttacca tggcggtagc gctagtacca ttatccccga atcaaagggc 1860
aatcaagcca gcatcatctt gacctggcac gctgcgacag gttgctacga caaagactat 1920
gagcaaagga agaagatgaa ggctactaca gaatcgccgc tacagcacag gcagtatcgt 1980
gttgtggcca ccgctctatc caagtactgc atgtacttgg tagcttatgt gcctcagctg 2040
ctgcccgggc agcagtccta cacaacatct gtctacaacg attttgtgcg ctcgccgtcc 2100
tacatcttgc agagtggtac ccagctgaaa gatgagctct gcgaggcggt ggtggaagat 2160
gaactgaggt ggaaggtgct agctgatttc tgggtcgaga tgttgctcta cctcgcgcct 2220
tctgacaatg tcacggctca catcgagcaa ctcgcacaag gtggagagtt catcacccat 2280
ctctgggctc tcctcttcca cgccggcatc ctgtaccggc cgtacaagga ggaggaggag 2340
ccagctgcag ccgggacagt gtga 2364
<210> 14
<211> 787
<212> PRT
<213>Rice (Oryza Sativa L)
<400> 14
Met Glu Gln His Val Glu Arg Asn Ala Ser Ser Ala His His Leu Arg
1 5 10 15
Asp Val Trp Leu Ser Ile Gly Gly Thr Val Val Arg Ile Glu Val Met
20 25 30
Ala Met Val Ala Ile Phe Leu Thr Phe Tyr Ala Val Thr Phe Gly Ser
35 40 45
Cys Arg Arg Trp Ser Ser Arg Trp Ile Ile Gln Lys Val Phe Leu Ala
50 55 60
Ala Asn Ala Leu Ser Val Ser Leu Gly Thr Tyr Ser Ile Gly Leu Met
65 70 75 80
Gln Ser Ser Leu Val Lys Ser Lys Met Tyr Pro Val Trp Ser Val Ser
85 90 95
Leu Leu Ala Leu Phe Ala Ser Ile Asp Ser Ile Asn Thr Phe Gly Leu
100 105 110
Asp Tyr Ser Gly Gln Phe Leu Lys Leu Leu Tyr Gln Leu Phe Leu Tyr
115 120 125
Phe Gly Tyr Val Leu Val Ile Ser Ile Ser Gly Asn Thr Asn Asn Gly
130 135 140
Met Val Ala Ala Ala Ile Gly Met Leu Cys Ser Val Thr Leu Cys Lys
145 150 155 160
Gly Phe His Arg Leu Met Ala Tyr Val Leu Pro Thr Arg Leu Arg Ser
165 170 175
Met Asn Lys Glu Ile Ala Asp Arg Met Glu Val Val Lys Gly Gly Pro
180 185 190
Asn Phe Asn Ala Ala Ile Met Gln Gly Tyr Gln Tyr Leu Val Asp Lys
195 200 205
Thr Tyr Tyr Pro Ser Ile Thr Leu Glu Glu Leu Trp Lys Pro Asp Asn
210 215 220
Gly Met Asp Lys Leu Gly Thr Asp Ala Asp Ala Tyr Lys Asp Ile Cys
225 230 235 240
Leu Ser Phe Ser Leu Cys His Leu Leu Gln Arg Arg Phe Phe Gly Phe
245 250 255
Ser Cys Ala Glu Ser Asp Arg Pro Glu Thr Arg Ser Phe Val Leu Glu
260 265 270
Gly Leu Leu Gln Pro Val Ala Ser Ser Ser Ser Ser Ser Gln Gly Thr
275 280 285
Gly Gly Ser Arg Gly Asn Tyr Glu Arg Ala Phe Lys Val Ile Glu Val
290 295 300
Glu Leu Ala Phe Met Tyr Asp Tyr Ile Tyr Ser Ser Asn Ala Phe Val
305 310 315 320
His Tyr Tyr Glu Ala Gly Ala Cys Thr Ala Trp Ala Ile Ala Ser Ile
325 330 335
Leu Thr Thr Cys Phe Leu Cys Val Ala Val Ala Leu Gln Gln Lys Glu
340 345 350
Ala Ile Ile Ala Ser Ser Ala Val Thr Met Leu Asp Ser Glu Val Ile
355 360 365
Ser Thr Thr Met Thr Pro Asp Ile Val Ile Thr Leu Val Ile Leu Val
370 375 380
Ser Leu Ala Ser Leu Gln Val Leu Gln Leu Val Asn Ser Leu Ser Ser
385 390 395 400
Asn Trp Ala Arg Val Ser Leu Val Cys His Leu Phe Arg Arg Asn Ala
405 410 415
Asn Asn Leu Glu Pro Ser Met Gly Leu Lys Leu Arg Leu Phe Leu Ser
420 425 430
Arg Ile Lys Leu Leu Asp Lys Tyr Gln Trp Gln Asn Lys Leu Gly Gln
435 440 445
Tyr Glu Ser Ser Ser Trp Leu Lys Lys Ala Leu Gly Phe Val Ser Phe
450 455 460
Lys Ile Gly Gln Cys Cys Tyr Lys Pro Val Phe His Phe Met Leu Tyr
465 470 475 480
Cys Val Arg Cys Met Met Arg Trp Pro Thr Ile Cys Cys Pro Cys Tyr
485 490 495
Val Val Cys Ser Pro Leu Cys Asn Trp Val Phe Lys Leu Gln Pro Ser
500 505 510
Trp Gln Leu Leu Pro Gly Pro Cys Trp Val Ile Tyr Ser Ala Leu Tyr
515 520 525
Lys Leu Phe Gly Leu Gln Tyr Ile Gln Gln Val Leu Thr Asp Met Leu
530 535 540
Gly Ile Ser Thr Ala Ser Ser Ile Gln Leu Pro Ile Glu Val Lys Ser
545 550 555 560
Thr Val Ile Asp Ala Leu Val Gly Ile Leu Met Pro Arg Pro Asn His
565 570 575
Asp Ser Val Val Leu Ser Ser Gly Ser Thr Ser Leu Ala Lys Asn Gly
580 585 590
Leu Gln Asp Lys Phe Ile Arg Gln Tyr Ala Ser Thr Ser Tyr His Gly
595 600 605
Gly Ser Ala Ser Thr Ile Ile Pro Glu Ser Lys Gly Asn Gln Ala Ser
610 615 620
Ile Ile Leu Thr Trp His Ala Ala Thr Gly Cys Tyr Asp Lys Asp Tyr
625 630 635 640
Glu Gln Arg Lys Lys Met Lys Ala Thr Thr Glu Ser Pro Leu Gln His
645 650 655
Arg Gln Tyr Arg Val Val Ala Thr Ala Leu Ser Lys Tyr Cys Met Tyr
660 665 670
Leu Val Ala Tyr Val Pro Gln Leu Leu Pro Gly Gln Gln Ser Tyr Thr
675 680 685
Thr Ser Val Tyr Asn Asp Phe Val Arg Ser Pro Ser Tyr Ile Leu Gln
690 695 700
Ser Gly Thr Gln Leu Lys Asp Glu Leu Cys Glu Ala Val Val Glu Asp
705 710 715 720
Glu Leu Arg Trp Lys Val Leu Ala Asp Phe Trp Val Glu Met Leu Leu
725 730 735
Tyr Leu Ala Pro Ser Asp Asn Val Thr Ala His Ile Glu Gln Leu Ala
740 745 750
Gln Gly Gly Glu Phe Ile Thr His Leu Trp Ala Leu Leu Phe His Ala
755 760 765
Gly Ile Leu Tyr Arg Pro Tyr Lys Glu Glu Glu Glu Pro Ala Ala Ala
770 775 780
Gly Thr Val
785
<210> 15
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 15
atggagcaac atgtggaacg 20
<210> 16
<211> 19
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 16
tcacactgtc ccggctgca 19
<210> 17
<211> 29
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 17
gcctctagaa tggagcaaca tgtggaacg 29
<210> 18
<211> 28
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 18
gccgtcgact cacactgtcc cggctgca 28
<210> 19
<211> 22
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 19
gggctgatgc agtcatcgct gg 22
<210> 20
<211> 20
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 20
ctccgcagcc tggtcgggag 20
<210> 21
<211> 21
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 21
accacttcga ccgccactac t 21
<210> 22
<211> 19
<212> DNA
<213>Artificial sequence (Artificial Sequence)
<400> 22
acgcctaagc ctgctggtt 19
<210> 23
<211> 2364
<212> DNA
<213>Rice (Oryza Sativa L)
<400> 23
atggagcaac atgtggaacg gaatgcatca agcgcacacc acctgagaga tgtgtggctg 60
agcattggag ggacggtggt ccgcattgag gtgatggcaa tggtggccat ctttctcacc 120
ttctacgccg tcaccttcgg atcctgccgc cgctggtcta gccgatggat catccaaaag 180
gttttcctgg ctgcgaatgc gctgtctgtg tccctaggaa catacagcat agggctgatg 240
cagtcatcgc tggtgaagag caagatgtac cctgtgtggt cggtgtcatt gcttgccctc 300
tttgctagca ttgactccat caacacattc ggcctcgact atagtggcca attcctgaag 360
ctgttgtatc agctcttcct ctactttggg tatgtgcttg tgatcagcat ctccggcaat 420
accaataatg gcatggtggc cgctgcgatt ggcatgcttt gcagcgtcac cctatgtaag 480
ggattccata ggctgatggc ctatgtgctc ccgaccaggc tgcggagcat gaacaaagag 540
attgcagatc gtatggaagt ggtgaagggg gggccaaact ttaacgcggc catcatgcaa 600
gggtatcaat acctggttga taagacttat tatccatcca taaccctgga agaattgtgg 660
aagcctgaca atggcatgga caaactagga acagatgctg atgcctacaa ggacatatgc 720
ctctctttct ccctctgtca cctgctacag cgacgcttct ttggttttag ttgtgctgag 780
tctgaccggc cagagacccg tagctttgtg ttggaaggct tgcttcaacc tgtagcttca 840
agtagttcat catcacaagg tactggtggt tctcgtggta actacgagag ggccttcaag 900
gtgattgagg ttgagctagc cttcatgtac gactacatct acagcagcaa cgccttcgtc 960
cattactatg aagctggagc ttgcaccgcc tgggcaatag cttcaatcct gaccacatgt 1020
tttctatgtg tggctgttgc cctacagcaa aaagaagcca ttattgcatc atctgcggtc 1080
actatgttgg actcggaggt aattagcaca acgatgacac cagacattgt catcaccctt 1140
gtcatactgg tgtcccttgc ttccctccaa gtgttgcagc tggtgaactc cttgtcgtcc 1200
aattgggcaa gagtctccct tgtctgtcat cttttcaaga gaaatgccaa caacctggag 1260
ccatcgatgg gcttgaagct gaggttgttc ctttccagga tcaaactatt ggacaaatac 1320
cagtggcaga acaagcttgg gcagtacgag agctcatcgt ggctcaagaa agctctaggg 1380
tttgtcagct tcaagatagg acaatgttgc tataagccag tgtttcattt catgctctac 1440
tgcgtgcggt gcatgatgcg atggccaacc atttgctgcc catgctatgt ggtgtgctct 1500
ccgctctgca actgggtgtt caaattgcag cctagttggc agttgctccc aggcccttgt 1560
tgggtgatat actccgccct ctacaagttg tttggcctgc agtacattca acaggtgctg 1620
acagacatgt tgggcattag cactgcaagc tccattcagc tgcccattga ggtgaagagt 1680
accgtgattg atgccctcgt tgggatcctc atgcccaggc ccaaccatga cagtgtcgtg 1740
ttgtccagtg gatcaacatc gctagccaag aatggcttgc aggacaagtt cataaggcag 1800
tacgcttcga caagttacca tggcggtagc gctagtacca ttatccccga atcaaagggc 1860
aatcaagcca gcatcatctt gacctggcac gctgcgacag gttgctacga caaagactat 1920
gagcaaagga agaagatgaa ggctactaca gaatcgccgc tacagcacag gcagtatcgt 1980
gttgtggcca ccgctctatc caagtactgc atgtacttgg tagcttatgt gcctcagctg 2040
ctgcccgggc agcagtccta cacaacatct gtctacaacg attttgtgcg ctcgccgtcc 2100
tacatcttgc agagtggtac ccagctgaaa gatgagctct gcgaggcggt ggtggaagat 2160
gaactgaggt ggaaggtgct agctgatttc tgggtcgaga tgttgctcta cctcgcgcct 2220
tctgacaatg tcacggctca catcgagcaa ctcgcacaag gtggagagtt catcacccat 2280
ctctgggctc tcctcttcca cgccggcatc ctgtaccggc cgtacaagga ggaggaggag 2340
ccagctgcag ccgggacagt gtga 2364
<210> 24
<211> 787
<212> PRT
<213>Rice (Oryza Sativa L)
<400> 24
Met Glu Gln His Val Glu Arg Asn Ala Ser Ser Ala His His Leu Arg
1 5 10 15
Asp Val Trp Leu Ser Ile Gly Gly Thr Val Val Arg Ile Glu Val Met
20 25 30
Ala Met Val Ala Ile Phe Leu Thr Phe Tyr Ala Val Thr Phe Gly Ser
35 40 45
Cys Arg Arg Trp Ser Ser Arg Trp Ile Ile Gln Lys Val Phe Leu Ala
50 55 60
Ala Asn Ala Leu Ser Val Ser Leu Gly Thr Tyr Ser Ile Gly Leu Met
65 70 75 80
Gln Ser Ser Leu Val Lys Ser Lys Met Tyr Pro Val Trp Ser Val Ser
85 90 95
Leu Leu Ala Leu Phe Ala Ser Ile Asp Ser Ile Asn Thr Phe Gly Leu
100 105 110
Asp Tyr Ser Gly Gln Phe Leu Lys Leu Leu Tyr Gln Leu Phe Leu Tyr
115 120 125
Phe Gly Tyr Val Leu Val Ile Ser Ile Ser Gly Asn Thr Asn Asn Gly
130 135 140
Met Val Ala Ala Ala Ile Gly Met Leu Cys Ser Val Thr Leu Cys Lys
145 150 155 160
Gly Phe His Arg Leu Met Ala Tyr Val Leu Pro Thr Arg Leu Arg Ser
165 170 175
Met Asn Lys Glu Ile Ala Asp Arg Met Glu Val Val Lys Gly Gly Pro
180 185 190
Asn Phe Asn Ala Ala Ile Met Gln Gly Tyr Gln Tyr Leu Val Asp Lys
195 200 205
Thr Tyr Tyr Pro Ser Ile Thr Leu Glu Glu Leu Trp Lys Pro Asp Asn
210 215 220
Gly Met Asp Lys Leu Gly Thr Asp Ala Asp Ala Tyr Lys Asp Ile Cys
225 230 235 240
Leu Ser Phe Ser Leu Cys His Leu Leu Gln Arg Arg Phe Phe Gly Phe
245 250 255
Ser Cys Ala Glu Ser Asp Arg Pro Glu Thr Arg Ser Phe Val Leu Glu
260 265 270
Gly Leu Leu Gln Pro Val Ala Ser Ser Ser Ser Ser Ser Gln Gly Thr
275 280 285
Gly Gly Ser Arg Gly Asn Tyr Glu Arg Ala Phe Lys Val Ile Glu Val
290 295 300
Glu Leu Ala Phe Met Tyr Asp Tyr Ile Tyr Ser Ser Asn Ala Phe Val
305 310 315 320
His Tyr Tyr Glu Ala Gly Ala Cys Thr Ala Trp Ala Ile Ala Ser Ile
325 330 335
Leu Thr Thr Cys Phe Leu Cys Val Ala Val Ala Leu Gln Gln Lys Glu
340 345 350
Ala Ile Ile Ala Ser Ser Ala Val Thr Met Leu Asp Ser Glu Val Ile
355 360 365
Ser Thr Thr Met Thr Pro Asp Ile Val Ile Thr Leu Val Ile Leu Val
370 375 380
Ser Leu Ala Ser Leu Gln Val Leu Gln Leu Val Asn Ser Leu Ser Ser
385 390 395 400
Asn Trp Ala Arg Val Ser Leu Val Cys His Leu Phe Lys Arg Asn Ala
405 410 415
Asn Asn Leu Glu Pro Ser Met Gly Leu Lys Leu Arg Leu Phe Leu Ser
420 425 430
Arg Ile Lys Leu Leu Asp Lys Tyr Gln Trp Gln Asn Lys Leu Gly Gln
435 440 445
Tyr Glu Ser Ser Ser Trp Leu Lys Lys Ala Leu Gly Phe Val Ser Phe
450 455 460
Lys Ile Gly Gln Cys Cys Tyr Lys Pro Val Phe His Phe Met Leu Tyr
465 470 475 480
Cys Val Arg Cys Met Met Arg Trp Pro Thr Ile Cys Cys Pro Cys Tyr
485 490 495
Val Val Cys Ser Pro Leu Cys Asn Trp Val Phe Lys Leu Gln Pro Ser
500 505 510
Trp Gln Leu Leu Pro Gly Pro Cys Trp Val Ile Tyr Ser Ala Leu Tyr
515 520 525
Lys Leu Phe Gly Leu Gln Tyr Ile Gln Gln Val Leu Thr Asp Met Leu
530 535 540
Gly Ile Ser Thr Ala Ser Ser Ile Gln Leu Pro Ile Glu Val Lys Ser
545 550 555 560
Thr Val Ile Asp Ala Leu Val Gly Ile Leu Met Pro Arg Pro Asn His
565 570 575
Asp Ser Val Val Leu Ser Ser Gly Ser Thr Ser Leu Ala Lys Asn Gly
580 585 590
Leu Gln Asp Lys Phe Ile Arg Gln Tyr Ala Ser Thr Ser Tyr His Gly
595 600 605
Gly Ser Ala Ser Thr Ile Ile Pro Glu Ser Lys Gly Asn Gln Ala Ser
610 615 620
Ile Ile Leu Thr Trp His Ala Ala Thr Gly Cys Tyr Asp Lys Asp Tyr
625 630 635 640
Glu Gln Arg Lys Lys Met Lys Ala Thr Thr Glu Ser Pro Leu Gln His
645 650 655
Arg Gln Tyr Arg Val Val Ala Thr Ala Leu Ser Lys Tyr Cys Met Tyr
660 665 670
Leu Val Ala Tyr Val Pro Gln Leu Leu Pro Gly Gln Gln Ser Tyr Thr
675 680 685
Thr Ser Val Tyr Asn Asp Phe Val Arg Ser Pro Ser Tyr Ile Leu Gln
690 695 700
Ser Gly Thr Gln Leu Lys Asp Glu Leu Cys Glu Ala Val Val Glu Asp
705 710 715 720
Glu Leu Arg Trp Lys Val Leu Ala Asp Phe Trp Val Glu Met Leu Leu
725 730 735
Tyr Leu Ala Pro Ser Asp Asn Val Thr Ala His Ile Glu Gln Leu Ala
740 745 750
Gln Gly Gly Glu Phe Ile Thr His Leu Trp Ala Leu Leu Phe His Ala
755 760 765
Gly Ile Leu Tyr Arg Pro Tyr Lys Glu Glu Glu Glu Pro Ala Ala Ala
770 775 780
Gly Thr Val
785

Claims (6)

1. rice leaf sugar accumulation related gene SAC1, it is characterised in that：The rice leaf sugar accumulation related gene SAC1's Nucleotide sequence is as shown in SEQ ID No.13.
2. rice leaf sugar accumulation related gene SAC1 according to claim 1, it is characterised in that：The Rice Leaf piece candy product The amino acid sequence of tired related gene SAC1 codings is as shown in SEQ ID No.14.
3. rice leaf sugar accumulation gene SAC1 mutators, it is characterised in that：The nucleotide sequence of the SAC1 mutators As shown in SEQ ID No.23.
4. rice leaf sugar accumulation gene SAC1 mutators according to claim 3, it is characterised in that：The SAC1 mutation The amino acid sequence of gene code is as shown in SEQ ID No.24.
5. application of the SAC1 mutators of claim 3 or 4 in the molecular breeding of rice leaf sugar accumulation character.
6. application according to claim 5, it is characterised in that：The kind of the rice is extensive No. 10 of red silk.
CN201711463928.3A 2017-12-28 2017-12-28 Rice leaf sugar accumulation related gene SAC1 and its SAC1 mutator and application Pending CN107893082A (en)

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108504772A (en) * 2018-06-05 2018-09-07 浙江农林大学 The molecular labeling of rice premature gene and application
CN111154769A (en) * 2020-01-21 2020-05-15 西南大学 Rice leaf sugar accumulation gene LSA1, protein coded by same and application thereof
CN114262710A (en) * 2021-12-31 2022-04-01 西南大学 Rice plasmodesmata gene and mutant gene, coded protein and application thereof

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
MUTATION OF THE OSSAC1 GENE, WHICH ENCODES AN ENDOPLASMIC RETICU: "Mutation of the OsSAC1 Gene, which Encodes an Endoplasmic Reticulum Protein with an Unknown Function, Causes Sugar Accumulation in Rice Leaves", 《PLANT CELL PHYSIOL.》 *
SASAKI,T. ET AL.: "GenBank：AP005486.3，Oryza sativa Japonica Group genomic DNA,chromosome 7,BAC clone:OJ1138_B05", 《GENBANK》 *

Cited By (6)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108504772A (en) * 2018-06-05 2018-09-07 浙江农林大学 The molecular labeling of rice premature gene and application
CN108504772B (en) * 2018-06-05 2021-06-11 浙江农林大学 Molecular marker of rice premature senility gene and application thereof
CN111154769A (en) * 2020-01-21 2020-05-15 西南大学 Rice leaf sugar accumulation gene LSA1, protein coded by same and application thereof
CN111154769B (en) * 2020-01-21 2022-06-10 西南大学 Rice leaf sugar accumulation gene LSA1, protein coded by same and application thereof
CN114262710A (en) * 2021-12-31 2022-04-01 西南大学 Rice plasmodesmata gene and mutant gene, coded protein and application thereof
CN114262710B (en) * 2021-12-31 2023-10-31 西南大学 Rice plasmodesmata gene, mutant gene thereof, coded protein and application

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