CN108739351A - The method for formulating ornamental type rice germplasm using two kinds of zinc finger protein genes of rice - Google Patents

The method for formulating ornamental type rice germplasm using two kinds of zinc finger protein genes of rice Download PDF

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CN108739351A
CN108739351A CN201810377397.4A CN201810377397A CN108739351A CN 108739351 A CN108739351 A CN 108739351A CN 201810377397 A CN201810377397 A CN 201810377397A CN 108739351 A CN108739351 A CN 108739351A
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rice
osjag
germplasm
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zinc finger
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段远霖
吴为人
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Fujian Agriculture and Forestry University
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    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/156—Polymorphic or mutational markers

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Abstract

Biotechnology of the present invention relates to the use of the method for two kinds of zinc finger protein gene initiative ornamental type rice germplasms of rice.With two kinds of protein gene containing Zinc finger domain of riceOsJAGWithDLMutation allele be that both allele are imported into same Indica Rice germplasm by raw material by hybridization and Marker-assisted selection, having formulated a kind of can stablize heredity, the high rice germplasm of ornamental value.The flower type structure of the germplasm is special, its four-wheel floral organ former base invariable number, but development direction occurs obviously to change, " Hua Zhongsui " that each Spikelet development is formed at the small ear by countless new anormogenesis, the periphery of these small ears is wrapped up by countless tiny green bran sheet organs, and centre is full of faint yellow, meat shape organ.Therefore, the rice germplasm formulated of the present invention has the characteristics that flower pattern is big and peculiar, the florescence is long, ornamental value is high, easy maintenance and breeding, can be used for cultivating ornamental type rice, with good application value and foreground.

Description

Method for creating ornamental rice germplasm by using two zinc finger protein genes of rice
Technical Field
The invention belongs to the technical field of plant biology, and particularly relates to a method for creating ornamental rice germplasm by two zinc finger protein genes of rice.
Background
The development of the floral organs of plants is a complex biological process that is regulated by the involvement of multiple genes. Over 20 years, a series of mutants in arabidopsis thaliana, petunia hybrida and snapdragon are utilized to carry out research, the research on the molecular genetic mechanism of the development of the floral organs of dicotyledonous plants is greatly progressed, and a plurality of plant floral organ development models determined by five functional genes of 'A, B, C, D, E' are provided. The five genes are mainly MADS-box and play a key role in the morphogenesis of plant organs, so the five genes are also called as 'flower organ characteristic' genes. As the most basic and key determinants of the formation of the floral organs, most of the floral organ characteristic genes are conserved in function, and the interaction and the cooperation of the floral organ characteristic genes determine the spatiotemporal sequence of the development of the floral organs.
Although floral organ signature genes play a critical role in the formation of floral organs in plants, they are not the only determinant. In recent years, it has been found that upstream regulatory genes, which are characteristic genes of flower organs, are involved in determining the formation of flower organs of plants by activating or inhibiting the expression of the characteristic genes of flower organs at different times, at different sites or in different regulatory pathways. Different from the common functions of characteristic genes of floral organs, the functions of the regulatory genes are obviously differentiated in monocotyledons and dicotyledons. The zinc finger protein is a transcription factor with finger-shaped structural domain which is commonly existed in plants, and has large quantity and wide distribution in the genome of eukaryotes such as plants. The existing research shows that part of zinc finger protein gene plays an important regulation role in the formation of floral organs of plants. Wherein, two genes containing zinc finger domainsOsJAGAndDL(YABBY gene containing zinc finger domain) was shown to play a key role in the morphogenesis of rice floral organs, especially sexual organs.OsJAGRegulation of floral organ (particularly stamen) development primarily by promoting expression of class B genes (Horigome et al, 2009; Xiao et al, 20)09, Duan et al, 2010); whileDLThe gene regulates the development of rice pistils by inhibiting the expression of class B genes in round 4 (Nagasawa et al, 2003; Yamaguchi et al, 2004). The research progresses, and lays a foundation for genetically improving the ear traits of the rice and creating a new rice germplasm.
Rice is an important grain crop, and the main purpose of people for planting rice is to obtain rice and provide staple food. The rice spike is an important agronomic character of rice and is composed of a plurality of small spikes. The spikelet is the basis of the formation of rice grains, the development condition of the spikelet directly influences the yield of rice, and any important gene related to the spikelet has functional mutation, so that the spikelet is abnormal in development and cannot bear fruit. With the improvement of living standard of people, people consume more and more ornamental plants such as flowers and the like, and the demand on the variety and the variety of the ornamental plants is higher and higher. The small flower of rice consists of the palea, the lemma, the blade, the stamen and the pistil from outside to inside in sequence. Normal rice has small flower and short flowering phase and has no ornamental value. However, the rice has the advantages of vigorous growth, beautiful plant shape, strong regeneration capacity, simple planting and maintenance, natural closeness of people to the rice and the like, and if the ears of the rice can be reformed, rice germplasm with large and fantastic flower types, long flower period and high ornamental value is developed, so the rice germplasm has good popularization and application prospects.
Disclosure of Invention
The invention aims to provide a method for creating ornamental rice by modifying the property of a rice spike part by using two zinc finger transcription factor genes of the rice. The two rice floral organ development regulating genes are respectively C2H2Type zinc finger protein geneOsJAGAnd YABBY family genes containing zinc finger domainsDLThe two genes can specifically regulate the development of the rice spikelet, and a new way is provided for improving the spike character of rice by using the two rice zinc finger protein genes so as to cultivate a new rice germplasm with high ornamental value.
The technical scheme of the invention is as follows:
the invention relates to two zinc finger protein genes for controlling the development of rice floral organsOsJAGAndDLthe nucleotide sequence of (5) was downloaded from the rice genome database (http:// www.ricedata.cn/gene/index. htm).
SaidOsJAGAllelic variants ofOsjagAndDLallelic variants ofdlAll are obtained from tissue culture progeny of indica rice variety Minghui 86. Wherein,Osjagis composed ofOsJAG8054bp of fragment of gene deletion;dlis composed ofDLThe intron 1 of the gene is inserted with a 5124bp retrotransposon gene, which results inDLThe gene is not expressed.
The double mutant produced by introducing the above two mutant alleles into the same rice plant by hybridizationOsjagdlThe development of the small spike is abnormal, and the formed new rice germplasm with large spike, peculiar structure, long flowering period, stable inheritance and extremely high ornamental value is provided.
Utilizing two zinc finger protein genesOsJAGAndDLallelic variant gene of (1)OsjagAnddlthe two alleles are introduced into the same indica rice with strong tillering and regeneration capacity by hybridization and molecular marker selection as raw materials to obtain germplasm with stable heredity and extremely high ornamental value.
The method specifically comprises the following steps:
(1) to carry withOsjagThe plant of (A) is used as female parent and is carrieddlAnd plants with normal pollen development are used as male parents for hybridization to obtain a hybrid F1;
(2) planting the F1 obtained in the step (1), and harvesting seeds of a single plant in a mature period to obtain F2 seeds;
(3) planting F2 obtained in step (2), thereby separatingOsjagAnddlin strains of two mutants, the molecular marker technology is used for quickly identifying the strains which are normal in fruit setting and carry the mutants simultaneouslyOsjagAnddlthe heterozygous genotype plants of (1) are harvested at the mature period to obtain the energyStably inherited double mutant heterozygous genotype germplasm;
(4) planting the seeds harvested in the step (3), and rapidly identifying double mutants from the seeds in the seedling stage by using a molecular marking technologyOsjagdl。
The allele of the mutant geneOsjagIs composed ofOsJAGGene deletion 8054bp fragment; the allele of the mutant genedlIs composed ofDLA5124 bp transposon gene is inserted into the 1 st intron of the gene.
As an improvement of the present invention, inOsJAGAndDLthe object of the present invention can also be achieved by an allele or a derivative thereof obtained by adding, substituting, inserting or deleting one or more nucleotides in the nucleotide sequence of the gene.
The invention has the advantages that: the normal rice floret has simple structure, consists of palea, pulp sheet, stamen and pistil from outside to inside, and has small flower type, short flowering period and no ornamental value. The invention uses two zinc finger protein genes of riceOsJAGAndDLthe mutant allele of the rice is taken as a raw material, the two alleles are quickly introduced into the same indica rice germplasm through hybridization and molecular marker selection, and the rice germplasm which has the advantages of stable inheritance, large flower type, peculiar structure, long flowering phase, high ornamental value, easy maintenance and propagation and the like is created, is used for cultivating ornamental rice, has low cultivation cost and high economic benefit, and has good application prospect.
Drawings
FIG. 1:OsJAGandDLdouble mutants of (2)OsjagdlThe phenotype of spikelets; wherein A: wild type, single mutantOsjagAnddlthe phenotype of spikelets; b: removing the wild spikelets after the inner husk and the outer husk; c: after removal of the inner and outer shellsOsjagSmall ears; d: after removal of the inner and outer shellsdlSmall ears; E-G: double mutants at 3 weeks post anthesisOsjagdlSmall ears of rice; h: double mutants at 3 weeks post anthesisOsjagdlOne small branch of spikelet; I-J: double mutants at 6 weeks post anthesisOsjagdlSmall ears of rice; scale bar =1mm (a-D), 2mm (E-J).
FIG. 2: retrotransposon gene insertionDLThe locus of the gene.
FIG. 3: primordial differentiation and developmental characteristics of the young ears of the double mutants at different developmental stages (numbers 1-6 and 7 in the figure indicate complete conversion of 6 stamen primordia from round 3 and 1 pistil primordia from round four into 7 other organ primordia). Scale bar =50 μm.
The specific implementation mode is as follows:
the present invention will be further described with reference to examples.
Example 1:Osjaganddlidentification and preservation of
OsjagIdentification and preservation of (1): rice mutants as shown in FIGS. 1A and 1COsjagObtained for the present inventors from a progeny population of young embryo cultures of the indica rice variety Minghui 86 (Duan et al, 2010). Mutants with significant floral organ dysplasia as shown in FIGS. 1A and 1COsjagThe inner and outer glumes of the 1 st round of the spikelet become narrow, thin and hard and cannot be closed, and the inner glumes are bent to be crescent; the white and fleshy pulp sheets in the 2 nd round are thinned and enlarged; the six stamens in round 3 are completely converted into pistil or pistil-stamen chimeras in the shape of meat; gynoecium dysplasia in round 4, increased stigma of the ovary and high sterility. Genetic analysis shows that the mutant character is controlled by recessive monogenic mutation. The sequencing result shows that the mutantOsJAGDeletion of 8054bp fragment (including 5) was detected from the gene’-Deletion 4492 bp before ATG of the start codon, 1416 bp of the gene region, and 3’-2146 bp after TGA for terminal stop codon). Therefore, the temperature of the molten metal is controlled,Osjagis a null allelic mutant (Duan et al, 2010).
Due to the fact thatOsjagThe pistil is highly sterile and the mutant character can only pass the heterozygous genotypeOsJAG/ OsjagStoring, and separating wild type (1/4), heterozygote (1/2) and hybrid from their selfing progenyOsjagMutant (1/4). According toOsjagThe length of the gene deletion sequence in the mutant is designed into 3 specific primers to form two pairs (primer 1+ primer 3, primer 2+ primer 3) which are mixed together, and 1 PCR reaction (denaturation at 94 ℃ for 30s, annealing at 55 ℃ for 30s and extension at 72 ℃ for 1 min) is utilized, so that the mutant can be obtainedOsjagThe three gene type plants are quickly identified from the seedlings of the hybrid selfing progeny, and the seeds of the hybrid gene type plants are harvested in the mature period. The sequences of the primers used and the results of the detection are shown in Table 1.
TABLE 1OsjagIdentification of three genotype plants of heterozygote selfing progeny
dlIdentification and preservation of (1): rice mutants as shown in FIGS. 1A and 1DdlObtained for the present inventors from progeny of a line cultured from immature embryos of indica rice variety Minghui 86. Taking immature seeds of indica rice variety Minghui 86 flowering 12 days later, sterilizing with 75% ethanol for 2 minutes, soaking with 2% sodium hypochlorite for 20min, rinsing with sterile water for 3-5 times, selecting young embryos, inoculating to NB culture medium (NB +2,4-D2 mg/L; pH 5.8-5.9), placing in a thermostat at 27 ℃ for dark culture for 15 days, and inducing callus to grow; transferring the grown callus into a new NB culture medium, and subculturing for 15 days; selecting yellow embryonic callus with smooth surface, transferring to differentiation culture medium (NB + KT 10 mg/L + NAA0.4 mg/L; pH 5.8-5.9), and differentiating into regeneration plant after 20-30 days; transferring the regenerated seedling with the height of 2-3cm to rooting culture medium (1/2 MS; pH 5.8-5.9) of 1/2MS, and culturing for 10-15 days; and transferring the regenerated plant after the seedling hardening treatment to a field for planting.
The mutants with obvious abnormal flower organs as shown in FIGS. 1A and 1D were found from the offspring of the young embryo tissue culture line of Minghui 86dl. The pistil of the mutant is completely degenerated and completedAll convert to multiple stamen organs of varying numbers, thus appearing as multiple stamens with no pistils. Sequencing results show that the mutant is shown in figure 2DLThe intron 1 of the gene is inserted with a 5124bp retrotransposon gene (the sequence is shown as SEQ ID NO. 1), which results inDLThe function is completely lost. Due to the fact thatdlThe pollen of the mutant can be bred but has no pistil and shows sterility, the mutant character can be preserved only by means of heterozygous genotype, and wild type (1/4), heterozygote (1/2) and heterozygote can be separated from its self-bred progenydlMutant (1/4). According todlIn the mutant, the retrotransposon sequence inserted in the gene is designed into 3 primers to form two pairs (primer 4+ primer 5, primer 4+ primer 6) to be mixed together, and 1 PCR reaction (denaturation at 94 ℃ for 30s, annealing at 55 ℃ for 30s and extension at 72 ℃ for 40 s) is utilized, so that the mutant can be obtaineddlWild-type and heterozygote plants are quickly identified from seedlings in the selfing progeny of the heterozygote, and seeds of the heterozygote gene plants are harvested at the mature stage. Can be directly identified according to the characteristics of leaf phenotypedlMutant plants. The sequences of the primers used and the results of the tests are shown in Table 2.
TABLE 2dlIdentification of hybrid inbred progeny wild type and hybrid plant genotype
Example 2: double mutantsOsjag dlCreation, rapid identification and preservation of
(1) To carry withOsjagThe plants of (4) as female parent (obtained in example 1)OsjagMutant hybrid) and carrierdlPlants with allelic mutation and normal pollen development (obtained in example 2)dlMutant heterozygote) as a male parent to obtain hybrid F1;
(2) planting the F1 obtained in the step (1), and harvesting seeds of a single plant in a mature period to obtain F2 seeds;
(3) planting F2 obtained in step (2), thereby separatingOsjagAnddlin strains of two mutants, the molecular marker technology is used for quickly identifying the strains which are normal in fruit setting and carry the mutants simultaneouslyOsjagAnddlthe heterozygous genotype plant is harvested in a mature period to obtain the double-mutation heterozygous genotype germplasm capable of being stably inherited;
(4) planting the seeds harvested in the step (3), and rapidly identifying double mutants from the seeds in the seedling stage by using a molecular marking technologyOsjagdl。
Due to two single mutantsOsjagAnddlcannot fruit normally, and their mutant characters can only be preserved by mutant heterozygote plants, respectively, with a ratio of 1/4 in their respective heterozygote selfing progenyOsjagMutants anddland occurs. Therefore, double mutants cannot be created by direct hybridization of two single mutants. Due to the fact thatdlDevelop normal stamens and pollen fertile, and hybrid plantsOsJAG/OsjagBoth stamens and pistils develop normally and become fertile. Thus, toOsJAG/OsjagAs the female parent anddlthe fertile pollen is hybridized to obtain F with normal development and fertility of stamen and pistil1Plant, selfed F thereof2Will isolate the double mutants as shown in FIGS. 1E-JOsjag dlAnd (5) plant growing. All singleOsjag dlThe spikelets are in a 'spike in flower' structure formed by a plurality of new abnormally developed spikelets, the peripheries of the spikelets are wrapped by a plurality of tiny green lemma sheet-shaped organs, and the middle parts of the spikelets are filled with light yellow and fleshy organs; under the condition of proper light-temperature environment,Osjag dlthe florescence of the flower can be more than 40 days. The observation of the scanning electron microscope shown in FIG. 3 shows that the double mutantsOsjag dlThe number of primordia of each round of floral organs of the phenotype of the spikelet is unchanged, but the development directions of four rounds of floral organs are obviously changed, and the phenotype and two single mutants with specific structure are finally developedOsjagAnddlcompletely different spikelets.
Since both mutants are from the same parent Minghui 86,Osjag dlthe mutant character of the double mutant can be stably inherited. Therefore, it is only necessary to select a plant containing two heterozygous genotypes at the same time, harvest the seeds of the double mutant heterozygous genotype plant at the mature stage to preserve the characters of the double mutant, and separate the double mutant from the population of the selfed progeny thereofOsjagdlAnd (5) plant growing. According toOsjagAnddlthe respective mutation characteristics can be obtained by 2 PCR methods using the primers in the above tables 1 and 2OsjagAnddlthe group of the self progeny of the double mutant heterozygote is quickly identifiedOsjagAnddldouble mutant heterozygote genotype plant and double mutant plant。Due to the fact thatOsjag dlDouble mutant plants anddlthe single mutant has the same characteristic of leaf draping, and when the double mutants are identified, the double mutants can be directly identified according to the phenotype of the leavesdlA mutant; then, the PCR method is used again to identifydlPresence or absence in the mutantOsjagThe genotype of the mutant.
TABLE 3OsjagAnddlidentification of 7 genotypes of selfing progeny of double-mutation heterozygote
Created by the inventionOsjagdlThe germplasm has high ornamental value and can be used for cultivating ornamental rice. The above description is only a few specific embodiments of the present invention, and it should be noted that all modifications that can be directly derived or suggested from the disclosure of the present invention by those skilled in the art are deemed to be within the scope of the present invention.
SEQUENCE LISTING
<110> Fujian agriculture and forestry university
<120> method for creating ornamental rice germplasm by using two zinc finger protein genes of rice
<130>7
<160>7
<170>PatentIn version 3.3
<210>1
<211>5124
<212>DNA
<213> Artificial sequence
<400>1
tgttggtgta aataagtgat gttcctagga cccatttgta aatgttaaga gaatagaaag 60
gaggagatgt gggaacaggg tagaaaacct gaaaaaaccc caaattagac aatatggtat 120
cagagccgag ctcctggcga ccagcggcgg caaaacccta gctggcggcg gctgcggcgg 180
cggcggctga gctgcggcgg cggcggcggc gctcgtgccg gagcaggacg aagtggaaga 240
cggtgtggag gcgaccgctg ggactcgcgg ggtggcggcg cgacctacgg cgacagcggc 300
tgagcggcgt gccggggtgg tcggccggtc acgcggccgt ggcgtggggg cgacggcgca 360
ggagtggtgg cggagctgcg accaggtgag ccggctggcc gctgttgctt gctgttgcta 420
cccgccgcgg ctggttgctg cgccggggct ggttgctgcg ccggcacgga ggacgacaag 480
aagggcaccg gctgctgttt gctgctgctg ttgcttgctg tttgctgctg ctgtggagag 540
aagggactgc tgtttgattg ctgtttgctg ctgctattgc ttgtttggtg gactgctgct 600
gtggagagaa gggaaaagag aggagaaggt tgctgttgct tgttgttggt agtgtcttgt 660
gtgctgaaaa attacaggag cagagagcag tagcgggtga ggactgagga agcttagaag 720
gggggagggg aaatgtcttc aggggttgga aaggaggttg tagatgcctc ctcggtggtt 780
actaatggtg atatgaaaga tttgctagag aatttgttga agatgggcat gataggtccg 840
aagagtgtag cagggggata cgaactgaaa ctagaattga tgccaaatga gttgaggttg 900
gaagggagca aaaattattt gagttggtgt aggagagccc aattaatgtt gagagcaaag 960
ggggtggatc attttttgca agagagttgt gaagagcctt ctgataagga gagccaagca 1020
tggagaacgt ggaatacaac aaattccact gtggtatctt ggttgatgac ctcggtggct 1080
ccttctattg gtaggatgat agagactatt cagaatgcag caattgtgtg gaagacattg 1140
agcaacatgt attcaggaga gggaaatgta atgatgatgg ttgaggctca gaacaaagta 1200
gaaaatttga agcaagaagg gagaacagtc caggagtacg ctagtgagtt gcagcaatta 1260
tgggcagatc ttgatcacta tgatcctcta cagttgaggc atgaggagga cattgtgatt 1320
ggaaataaat ggatgcagag gcgaagagtc attcatttct taaaggggtt gaacaaggag 1380
tttgaggaca gaagagcggc tatgttctac caggctacat tgcccactat ggaagaggct 1440
atttctgcta tggtgcagga ggagatgagg ttgaaactga tgagaggtac aaatcctaca 1500
agatcagcat acactgtggc tgataataga gaatgctaca actgtggaca agtgggtcat 1560
gtgagctaca attgtcccac tcctcggaac attggcggta gggggttgat tcgaggaggg 1620
tatggtggat ttggtggaac ccgtggtgga tttggaggag accgtggtga ctttgggggg 1680
aaccacagtg gtagaggagg tcgtggtgga gatcgtggtg gtggaggtcg tggtaggggc 1740
gaggtgctcc tcaggccatg tagccataga ggagggtaaa gctattacct taacaggtga 1800
acaggtgaca cagtgggagg aatggcagaa gaacaagatc aatgagagct ccaacaccac 1860
cactcacttt ggtaacttcg ccaactacgc tcaagtgggc gaaggtattc aggcacaggc 1920
acttgcatct acatacagac atcctataga ttggatcata gactcaggag catcaaagca 1980
tttcacagga ttgcataata ccttcacatc atacacccct tatgtccact ctgaaactat 2040
ccaaatcgct gatggtacat ccaaacctat tcatggtata gggtcagtag agtgcacatc 2100
atcaataaac ttatcctctg acttgcatgt tccttccttt ccagtaaacc ttctctcagt 2160
tagctcagct attgaccaac tcaagtgcat tgttgtattt gatgagaact ctcatctgtt 2220
ccaggagaag gggactggga ggaggattgg gactagagtc aggtgtgatg ggttgtggta 2280
catcaatcat gaggaactgg ggctagctgc ggtggttgga aatgttgaga aggagatcag 2340
tttgcttcat tgtcagttag gacatccatc ttttgagatt ttgagtaagt tgtatctaga 2400
tctctttagt aaagtggata agcatagatt ggtgtgtgat gcttgtgagc ttggaaaaca 2460
tactcggtct acatatgttg ggattggtct tcgtaactgt gagcctttca tattaataca 2520
ttctgatgtt tggggaccat gcccagttac ttctgtgagt ggttttaagt ggtttgtcac 2580
ctttattgat tgccacactc gtatgacttg gatttatatg cttaagcaca atagtgaggt 2640
ccttcgttgc tttcaggact ttcataaatt agtgacaact cagtttgatg caaaagttaa 2700
gattattcgg actgataatg gaacagaata tatcaataat gaatttgtgt catatatctc 2760
agatgagggg attattcacc agactagtac ccctcctcaa aatggtgtag ctgagagaaa 2820
gaatcgacat ttgctagaag tggcaagatc tttaatgttc cagatgaatg tgccaaagta 2880
cctatggagt gaggctgtga tgacagctgc atatcttatc aatcgcatgc cttctagaat 2940
acttggtatg aagtcccctg ttgaactttt attgggtaag cgagagttca aggtccctcc 3000
aaaggtgttt ggttgtgttt gctttgtgcg agatcatcga ccttctattg gcaagttgga 3060
tcctcatgca gtaaagtgtg tgtttgttgg ctatgcttca agtcagaagg ggtataaatg 3120
ttgggatccc attgggagaa gactgtttgt gagtatggat gtgacatttc gtgagtttga 3180
gccctattat aaaagtaaag gagatcttga ccaatttctt gaggaattct cgactgtcat 3240
ggaggttgat agtcgagagg gggagataga gagaggtgac acacatagga aaaatgttag 3300
tgacaagaat ggggagacag tggtggttgg atcaatacca tgctctattg acaatgcaag 3360
taaagaagca gtggaagtta ttggagatac acaagacaag gatagagaaa tggttcttca 3420
tgaggaggat ggagaagatg gagaagatga agaagtggtt gttgggacaa ttccatgtcc 3480
tatggagcga gccgaaaagg tgaaacaaaa ggatgtgcta gtctaccaaa ggaggcggtt 3540
tgatagtcag ggggagaaaa gaaaagggtt agtgcaaagt taagttgaag agttatcaca 3600
tccagaatgc ccagtccctg aatcctctca gaacctctct cctacagtct cattggctcc 3660
ccttgagata attggtaata ctcctcccat tcttgaacaa gtagaattac ctcttgcaca 3720
gcgtagagag actagatcca atgctggtag acctcctata cgtcttggtt ttgagcatct 3780
tagttctatg catgacattg caaactatat cttatattct catgtttcac tggcatacaa 3840
aacattcatt gcatcattat agacagtgcc cataccaaag gattggaagt gtgcaaaaca 3900
agatccaagg tggaaagatg caatgagaga agagttgaat gcccttatga agaataagac 3960
ttgggagctt gtaaaacttc caccaggaaa aaaggcagtg ggatgtaagt gggtttttac 4020
agtgaagcaa actcctgaag ggaaggtgga taggtataaa gcaagattag tcgcaaaagg 4080
ctacagtcaa acatatggaa ttgactatga tgagacattt gcaccggtgg caaaaatggg 4140
tacagtgaga actttagtat cttgtgcagt gaactttggg tggcccctcc atcagcttga 4200
tgtgaaaaat gcatttcttc atggtgattt atatgaggag gtctacatgg agattcctcc 4260
aggatttggg aatagccaga cagttggaaa aatatgtaaa ctcaagaaat ccttgtatgg 4320
gttgaagcag tttccacgtg cgtggtttga aagattcagg tgcgcagtgt gtgatatggg 4380
gtattctcag tgtaatggag atcacacggt attctacaaa cataggggga catacatcac 4440
catcttggct gtgtatgttg atgatatagt gatcaccggt gatgatgtag aagaaatcag 4500
gtgcctgaag gaatggacac ctaaatgtag aaggatattg tgatgctgat tgggcgagta 4560
gtatggatga tagaaggtct acatcaggtt attgtgtgtt tgttggtggt aatcttgttt 4620
cttggcgaag caagaagcag gcaatggtgg ctcggtctac cgccgaagca gaatatagag 4680
ctatggcttt gagtttgagt gagatgctgt ggatgaggag cttgttgact gaattgcgag 4740
tgttaagaag tgatactgtg atgcttcatt gtgacaataa atcagccatc aacattggaa 4800
acaaccttgt tcaacatgat cgcactaagc atgtggagat tgataggttc tttattaagg 4860
agaagattga tagtggggta cttatgctag agtatatcaa gtcatgtgaa caactagctg 4920
attgtctaac taaggggtta ggtcctagtg aaattcaatc aatatgtaac aagatgggta 4980
tgattgatat cttttaccca tcttgagggg gagtgttggt gtaaataagt gatgttccta 5040
ggacccattt gtaaatgtta agagaatata aaggaggaga cgtgggaaca ggggagtaac 5100
cctgaaaaac cccccaaaat taga 5124
<210>2
<211>20
<212>DNA
<213> Artificial sequence
<400>2
ttggatcatg gtgagggaga 20
<210>3
<211>19
<212>DNA
<213> Artificial sequence
<400>3
tggggggttg ggtatagta 19
<210>4
<211>19
<212>DNA
<213> Artificial sequence
<400>4
atggtgcaga gtgacgaca 19
<210>5
<211>20
<212>DNA
<213> Artificial sequence
<400>5
gatagctggg accaaatcca 20
<210>6
<211>21
<212>DNA
<213> Artificial sequence
<400>6
ggagaaaaag tagaggagaa c 21
<210>7
<211>20
<212>DNA
<213> Artificial sequence
<400>7
ctaccaacaa caagcaacag 20

Claims (4)

1. A method for creating ornamental rice germplasm by using two zinc finger protein genes of rice is characterized in that the two zinc finger protein genes are usedOsJAGAndDLallelic variant gene of (1)OsjagAnddlthe two alleles are introduced into the same indica rice with strong tillering and regenerating capability through hybridization and molecular marker selection as raw materials to obtain germplasm which can be stably inherited and has ornamental value.
2. The method for creating ornamental rice germplasm according to claim 1, wherein the method comprises the following steps:
(1) to carry withOsjagThe plant of (A) is used as female parent and is carrieddlAnd plants with normal pollen development are used as male parents for hybridization to obtain a hybrid F1;
(2) planting the F1 obtained in the step (1), and harvesting seeds of a single plant in a mature period to obtain F2 seeds;
(3) planting F2 obtained in step (2), thereby separatingOsjagAnddlin strains of two mutants, the molecular marker technology is used for quickly identifying the strains which are normal in fruit setting and carry the mutants simultaneouslyOsjagAnddlthe heterozygous genotype plant is harvested in a mature period to obtain the double-mutation heterozygous genotype germplasm capable of being stably inherited;
(4) planting the seeds harvested in the step (3), and rapidly identifying double mutants from the seeds in the seedling stage by using a molecular marking technologyOsjagdl。
3. The method for creating ornamental rice germplasm according to claim 1, wherein the allele of the zinc finger protein gene is a mutant of a rice plantOsjagIs composed ofOsJAG8054bp of fragment of gene deletion; the allele of the mutant genedlIs composed ofDLA5124 bp transposon gene is inserted into the 1 st intron of the gene.
4. The method according to claim 1, further comprising using as a starting material an allele or derivative resulting from addition, substitution, insertion or deletion of one or more nucleotides or the like in the sequences of the two genes.
CN201810377397.4A 2018-04-25 2018-04-25 The method for formulating ornamental type rice germplasm using two kinds of zinc finger protein genes of rice Pending CN108739351A (en)

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2010048398A2 (en) * 2008-10-22 2010-04-29 Cornell University Mutated eif4e sequences from potato which are useful in imparting virus resistance
CN106982733A (en) * 2017-02-20 2017-07-28 浙江大学 A kind of selection of pink blade rice varieties
CN107155873A (en) * 2017-07-17 2017-09-15 安徽省农业科学院水稻研究所 One kind makes the acarpous producing method for seed of hybrid paddy rice male parent

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2010048398A2 (en) * 2008-10-22 2010-04-29 Cornell University Mutated eif4e sequences from potato which are useful in imparting virus resistance
CN106982733A (en) * 2017-02-20 2017-07-28 浙江大学 A kind of selection of pink blade rice varieties
CN107155873A (en) * 2017-07-17 2017-09-15 安徽省农业科学院水稻研究所 One kind makes the acarpous producing method for seed of hybrid paddy rice male parent

Non-Patent Citations (3)

* Cited by examiner, † Cited by third party
Title
HAIFENG LI等: "The AGL6-1ike gene OsMADS6 regulates floral organ and meristem identities in rice", 《CELL RESEARCH》 *
YUANLIN DUAN等: "Molecular cloning and functional characterization of OsJAG gene based on a complete-deletion mutant in rice (Oryza sativa L.)", 《PLANT MOL BIOL》 *
郑莉莉: "dl/Osjag双突变体的形态特征与表达分析", 《中国优秀硕士学位论文全文数据库基础科学辑》 *

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Application publication date: 20181106