CN108018290A - Anthocyanin synthesis control gene and its application - Google Patents
Anthocyanin synthesis control gene and its application Download PDFInfo
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- CN108018290A CN108018290A CN201711269495.8A CN201711269495A CN108018290A CN 108018290 A CN108018290 A CN 108018290A CN 201711269495 A CN201711269495 A CN 201711269495A CN 108018290 A CN108018290 A CN 108018290A
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- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/415—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from plants
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- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/82—Vectors or expression systems specially adapted for eukaryotic hosts for plant cells, e.g. plant artificial chromosomes (PACs)
- C12N15/8241—Phenotypically and genetically modified plants via recombinant DNA technology
- C12N15/8242—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits
- C12N15/8243—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine
- C12N15/825—Phenotypically and genetically modified plants via recombinant DNA technology with non-agronomic quality (output) traits, e.g. for industrial processing; Value added, non-agronomic traits involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine involving pigment biosynthesis
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- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
- C12Q1/6895—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for plants, fungi or algae
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- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/13—Plant traits
Abstract
The invention discloses Anthocyanin to synthesize control gene and its application.Disclosed gene order is as shown in SEQ ID No.1.The insertion mutation of large fragment of the gene in the First Intron of white Chinese Cabbage there are a 3772bp size, its gDNA sequence are as shown in SEQ ID No.5.Mutation in introne in the gDNA nucleotide sequences of the gene devises Marker1, and Marker2, Marker8 tri- is to Specific PCR primers, and wherein Marker1 and Marker2 are codominant marker, and Marker8 marks for maternal instinct.By carrying out PCR amplification to the DNA of target Chinese cabbage, being identified through agarose gel electrophoresis, Marker1 and Marker2 energy Rapid identifications go out the strain of drinamyl heterozygosis and homozygous strain in segregating population, with reference to leaf-head character, Marker8 can also distinguish the drinamyl heterozygosis and homozygous strain of segregating population.
Description
Technical field
The invention belongs to vegetable breeding and molecular genetics field, and in particular to the base of control drinamyl Chinese cabbage Purpled traits
Cause, molecular labeling and its application.
Background technology
Chinese cabbage (Brassica rapa L.) belongs to Cruciferae Brassica genus Vegetable Crops of Brassica Chinese cabbage subspecies, is initiated by China
One of main vegetables crop, and one of vegetable crop of China's cultivated area maximum, with daily life health closely
It is related.It is orange, purple is big as the improvement of people's living standards, in recent years, requirement of the people to Chinese cabbage quality increasingly improves
Chinese cabbage breeding is paid attention to be subject to more and more domestic and foreign scholars and breeder.
Purple Chinese cabbage not only contributes to improve the anti-adversity ability of Chinese cabbage group itself since anthocyanidin content enriches,
And it is high for the nutritive value of human health, it has effects that anticancer, anti-oxidant and angiocardiopathy are subject to both at home and abroad
Extensive concern.But drinamyl Chinese cabbage leaf-head color needs just show after its balling maturation, and need in field
Cut leaf-head to observe by strain, operate not only very big but also time-consuming and laborious to the destructiveness of germplasm materials, greatly delay
Breeding process.Therefore clone controls the gene of Chinese cabbage purple leaf-head, studies its simple accurately detection method for innovation kind
Matter resource, quickening breeding paces seem particularly significant.
Applicant uses map based cloning and candidate gene analysis of gene differential expression method, with reference to the sequencing knot of Chinese cabbage
Fruit, the gene of screening and cloning control Chinese cabbage purple leaf-head simultaneously develop and Chinese cabbage leaf according to the sequence difference of the gene
The molecular labeling and dominant marker that ball purple gene isolates, to carry out, purple leaf-head Chinese cabbage forms molecule mechanism and molecule is auxiliary
Breeding is helped, accelerates breeding process and lays a good foundation.
The content of the invention
The present invention provides Anthocyanin to synthesize control gene.
Anthocyanin provided by the invention synthesis controls the sequence of gene to be：
(1)SEQ ID NO:Nucleotide sequence shown in 1；Or
(2)SEQ ID NO:Nucleotide sequence shown in 1 be substituted, lack and/or increase one or more nucleotide and
Express the nucleotide sequence of identical function protein；Or
(3) there is more than 90% homology with the nucleotide sequence of (1) or (2) and expresses the nucleosides of identical function protein
Acid sequence.
Present invention also offers expression cassette, expression vector, cloning vector or engineering bacteria, it includes the core for including the present invention
Acid.
The gene of the present invention synthesizes the application in anthocyanidin in plant, and the plant includes：Chinese cabbage, balling is not white
The various plants such as dish, arabidopsis.
Invention further provides the construction method of genetically modified plants, it utilizes genetic engineering means, the mistake in target plant
The gene of the present invention is expressed, screens positive transgenic plant.
Invention also provides with the relevant molecular labeling of Chinese cabbage anthocyanidin synthetic gene, it is characterised in that it is described
Insertion mutation, its gDNA sequence comprising a 3772bp fragment in molecular labeling is as shown in SEQ ID No.5.
Invention further provides the PCR detection kit for identifying drinamyl Chinese cabbage material, it is characterised in that the examination
Agent box contains the sense primer and anti-sense primer for being useful for molecular labeling described in amplification claim 5：
Sense primer：5’-ATGGAGGGTTCGTCCCAAG-3’
Anti-sense primer：5’-TCAAGTTCCAGTTTCTCCATCC-3’.
Present invention also offers the PCR detection kit for identifying drinamyl Chinese cabbage material, it is characterised in that the examination
Agent box contains the sense primer and anti-sense primer for being useful for molecular labeling described in amplification claim 5：
Marker1-F：5’-TGGTGTACTTTGATCCTTCGTG-3’
Marker1-R：5’-ACTGCATTCGCGTCTCCTAC-3’
Marker2-F：5’-CTTTTCTGCACGAACCCG-3’
Marker2-R：5’-ATTCGCGTCTCCTACTCCATAT-3’
Marker8-F：5’-AAAAGGTGCATGGACTGCT-3’
Marker8-R：5’-TGTTTTGGTGTCGATGAAGAAG-3’.
Application of the molecular labeling of the present invention in purple Chinese cabbage molecular mark.
Application of the molecular labeling of the present invention in purple Chinese cabbage material is identified.
In the present invention studies, the Specific PCR primers and detection method of inventor's design can not only be to Chinese Cabbages
The DNA extracted carries out PCR amplification, moreover it can be used to largely screens Chinese-cabbage Germplasm.The PCR primer is to utilize design of primers
Software Primer 5.0, is designed based on the sequence difference between the BrMYB2 genes and its allele of drinamyl Chinese cabbage
's.Chinese cabbage cultivar and germ plasm resource containing this gene can be quickly detected according to the presence or absence of amplified band and length size,
Its accuracy detected is high, easy to operate, of low cost.If containing this gene in a Chinese cabbage cultivar or germ plasm resource,
It is 5442bp or the specific fragment of 1665bp that can amplify length with above-mentioned primer, wherein big band is with white leaf-head
The plant of genetic background, small band are then the plant with purple leaf-head genetic background.
Present invention obtains Chinese cabbage control drinamyl leaf-head character (/ related to anthocyanidin synthesis) gene order and its
Quickly detect whether the detection method containing the gene and its allele.Heterozygosis can just be carried out in seedling stage by developing two pairs
The codominant marker of strain identification, and a pair of inclined maternal instinct that leaf-head Characters Identification heterozygosis strain is combined in the ball phase mark, and utilize
This method can quickly detect the Chinese cabbage plant containing the gene, its detection accuracy is high, easy to operate, of low cost.Make
Obtain and largely screen and identify that genotype is possibly realized in purple Chinese-cabbage Germplasm, mark property can also be used as in purple
Applied in heart cabbage hybrid Purity.The equipotential of the one group of control drinamyl Chinese cabbage Purpled traits obtained by the present invention
Gene (BrMYB2 and Brmyb2), the mechanism study available for drinamyl Chinese cabbage anthocyanidin synthetic molecules mechanism.
Brief description of the drawings
Fig. 1 is the BrMYB2/Brmyb2 Gene specific PCR primers of the invention amplification in cDNA.W, which is represented, to be contained
The Archon Chinese cabbage pcr amplification product of Brmyb2 genes, P represent the amplified production of the drinamyl Chinese cabbage containing BrMYB2 genes, M
Represent the amplified production of the purple tsai-tai parent containing BrMYB2 genes；M1 is DNA ladder.
Fig. 2 is BrMYB2/Brmyb2 genes amplification in gDNA, the common Chinese cabbage amplification containing BrMYB2 genes
Go out the band of a 1665bp, and the drinamyl Chinese cabbage containing Brmyb2 genes then amplifies the band of a 5442bp.W is represented
Archon Chinese cabbage pcr amplification product containing Brmyb2 genes, P represent the amplification production of the drinamyl Chinese cabbage containing BrMYB2 genes
Thing, M represent the amplified production of the purple tsai-tai parent containing BrMYB2 genes；M1 is DNA ladder；M2 is DNA ladder.
Fig. 3 is the codominant marker that Marker1 and Marker2 are amplified, and the Chinese cabbage containing BrMYB2 genes amplifies
The band (Marker1 306bp, Marker2 348bp) of one treaty 300bp, and the Chinese cabbage containing Brmyb2 genes is then expanded
Increase the band (Marker1 4081bp, Marker2 4123bp) for an about 4000bp.And heterozygosis strain two couple mark
The band of two kinds of sizes can be amplified.W represents the Archon Chinese cabbage pcr amplification product containing Brmyb2 genes, and P, which is represented, to be contained
There is the amplified production of the drinamyl Chinese cabbage of BrMYB2 genes, W+P represents the amplified production of heterozygosis drinamyl Chinese cabbage.
Fig. 4 is the dominant marker that Marker8 is amplified, and the Chinese cabbage containing Brmyb2 genes then amplifies a 972bp
Band, and the Chinese cabbage containing BrMYB2 genes is without special amplified production.If the Chinese cabbage with purple leaf-head character
The band can also be amplified, then it is heterozygosis strain to illustrate the plant；W represents the Archon Chinese cabbage PCR containing Brmyb2 genes and expands
Increase production thing, P represents the amplified production of the drinamyl Chinese cabbage containing BrMYB2 genes, and W+P represents the amplification of heterozygosis drinamyl Chinese cabbage
Product.
Embodiment
Following embodiments material therefor：
Drinamyl Chinese cabbage strain 11S96 (Duan Yanjiao, Zhang Lugang, He Qiong, Zhang Mingke, stone Jiang Chao drinamyl Chinese cabbage anthocyanidin
Characteristic of accumulation and related gene expression analysis [J] gardening journals, 2012Vol.39 (11):It is 2159-2167) by subspecies
Hybridization, and the strain that the various economical characters of inbreeding of more generation incubation are stablized.Inventor is under study for action by this strain and orange great Bai
(14S162 is selected more for individual plant selfing by orange Chinese cabbage ' S941-6 ' to dish " 14S162 ", and ' S941-6 ' is disclosed in ZL
The 2006 F1 colonies 10104743.9) produced for hybridization of female parent, then obtain corresponding F2 colonies by individual plant selfing.In F2
In colony, selection have purple, without purple each 10 plants build DNA ponds, using BSA methods full-length genome screen compact linkage molecule mark
Target gene, is as a result positioned on A7 chromosomes, one-step cloning of going forward side by side obtains the gene sequence of drinamyl Chinese cabbage by note
Row, as shown in SEQ ID No.1.Allelic sequences in white Chinese Cabbage, as shown in SEQ ID No.5.
Embodiment 1：The extraction of Chinese cabbage genome total serum IgE
1) the material liquid nitrogen grinding of cryopreservation is taken, the powder of about 50mg grindings, adds 1mL Bizol solution, acutely
Mix, stand 15min on ice；4 DEG C, 12000 × g centrifugation 15min, take supernatant, add 200 μ L chloroforms, cover tube cover, acutely shake
Swing, stand 15min on ice；4 DEG C, 12000 × g centrifugation 15min, careful Aspirate supernatant adds isometric pre- in another centrifuge tube
The isopropanol of cold (- 20 DEG C), turns upside down and mixes for several times, -20 DEG C of precipitation more than 2h；
2) in 4 DEG C, 10000 × g centrifuges 10min.Abandon supernatant, add 75% ethanol of 1mL (gone out without enzyme ddH2O is prepared,
Contain 0.1%DEPC, 40min sterilizings gained) wash precipitation twice；Supernatant is abandoned, RNA about 10min is air-dried on superclean bench, adds
50-100 μ L are without enzyme ddH2O dissolves；The genomic DNA in RNA is removed with RQDNase I (TaKaRa, Dalian), specific method is such as
Under：Following reacted constituent is continuously added in the centrifuge tube containing thick RNA：10 × Reaction Buffer10 μ L, RNA
Inhibitor (40U/ μ L) 1 μ L, DNase I (5U/ μ L) 6 μ L, add no enzyme ddH2O to 100 μ L of cumulative volume, mix, of short duration centrifugation,
37 DEG C of reaction 30min；
3) (24/1) 300 μ L of chloroform/isoamyl alcohol are added, are vigorously mixed vibration, add 300 μ L without enzyme ddH2O, it is fully mixed
It is even, 15min is placed on ice, is shaken once every a few minutes；4 DEG C, 12000rpm centrifugation 15min, careful Aspirate supernatant, is transferred to another
In centrifuge tube, the isopropanol of isometric precooling (- 20 DEG C), -20 DEG C of precipitation more than 2h are added；4 DEG C, 10000rpm centrifugations
10min recycling precipitations, abandon supernatant, and precipitation is washed twice with 75% ethanol (4 DEG C of placements)；The synthesis of superclean bench cDNA uses
One chain synthetic agent box (D6110A) of TaKaRa companies carries out, and concrete operations are as follows：
4) following reacted constituent is added in PCR pipe：About 5 μ g, Oligo (dT) 18 (50 μm of ol/L) of total serum IgE 1 μ L, dNTP
(10mmol/L) 1 μ L, no enzyme ddH2O supplies 10 μ L；5min is denatured in 65 DEG C, is placed in 2min on ice.Continue into above-mentioned PCR pipe
Add following ingredients：5 × RT Buffer, 4 μ L, RNase Inhibitor (40U/ μ L), 0.5 μ L, PrimeScript Rtase
(200U/uL) 1 μ L or M-M μ LV Reverse Transcriptase (5U/ μ L) 1 μ L, dd H2O 4.5 μ L, 20 μ L of cumulative volume,
Mixing slightly centrifuges, and 42 DEG C of warm bath 1h, 70 DEG C of 15min inactivate enzyme, are subsequently placed in 4 DEG C of coolings, stand-by.
Embodiment 2：The extraction of Chinese cabbage genome DNA
1) take 0.2g to remove the fresh leaflet tablet of Chinese cabbage of master pulse, fill in that (steel ball needs equipped with steel ball
75% alcohol washes are crossed) 2mL centrifuge tube in, be put into quick-frozen in liquid nitrogen, clayed into power using tissue grinder instrument；
2) CTAB extracting solution (CTABs of the 700 μ l through 65 DEG C of preheatings is added into centrifuge tube：2%, Tris-HCl (pH=8.0)：
100mmol/L, EDTA：20mmol/L, NaCl：1.4mol/L), the beta -mercaptoethanol of 10 μ L is added, it is rapid to mix；
3) then centrifuge tube is put into 65 DEG C of baking ovens, centre was shaken once at interval of 5-10min minutes, warm bath 45min；
4) centrifuge tube is taken out, adds isometric phenol：Chloroform：Isoamyl alcohol (25:24:1) mixture, after shaking up 15min,
10000r/min centrifuges 10min under room temperature；
5) upper phase (about 700 μ l) is transferred in another centrifuge tube, adds isometric chloroform and isoamyl alcohol mixing
Solution (chloroform：Isoamyl alcohol=24：1) 10min, is gently shaken up, 10000r/min centrifuges 10min under room temperature；
6) supernatant (about 500 μ l) is taken, adds the pre-cooled absolute ethyl alcohol of 2 times of volumes, gently mixing makes DNA unite, -20 DEG C
Under the conditions of precipitate 30min, 8000r/min centrifugations 5min under the conditions of 4 DEG C；
7) supernatant is abandoned, after the ethanol washing that 500 μ l mass percents of addition are 75% precipitates 2 times, sediment room temperature is dried；
8) add 500 μ l sterile distilled water dissolving DNA, add 0.29 μ l RNaseA (10 μ g/ μ l), after mixing slightly from
The heart, 37 DEG C of insulation 30min；
9) the 50 μ L of NaAc solution of 3mol/L and the absolute ethyl alcohol that 2 times of volumes are pre-cooled are added, gently mixing embraces DNA
, precipitate 30min under the conditions of -20 DEG C；
10) under the conditions of 4 DEG C, 8000r/min centrifugation 5min, abandon supernatant, it is clear to add the ethanol that mass percent is 75%
After washing precipitation 1~2 time, sediment room temperature is dried, add the μ l of 400 μ l~500 without enzyme ddH2O dissolvings use.
Embodiment 3：The PCR amplification of Chinese cabbage purple gene and genetic test
Using STb gene in embodiment 2 as masterplate, drinamyl Chinese cabbage purple gene is obtained respectively, the purple in purple tsai-tai parent
The detection of the gene of character and its allele and the gene in white leaf-head Chinese cabbage.
1) 20 μ l PCR reaction systems are：The template DNA of 50ng/ μ l is 2 μ l, 2 × Taq Master Mix 10 μ l, 10 μ
Each 1 μ l of upstream and downstream primer of M, reaction system is supplemented to 20 μ l with aseptic deionized water.
Primer sequence is：
Sense primer：5’-ATGGAGGGTTCGTCCCAAG-3’
Anti-sense primer：5’-TCAAGTTCCAGTTTCTCCATCC-3’
2) pcr amplification reaction carries out in PCR instrument
Temperature and reaction condition are：94 DEG C of pre-degeneration 3min；Then 94 DEG C of denaturation 30s, 58 DEG C of annealing 30s, 72 DEG C extend,
Totally 35 circulations；Last 72 DEG C re-extend 8min, 4 DEG C of preservations.Wherein 72 DEG C extension 4min are clone of the gene in gDNA
Used with detection, 72 DEG C of extension 1min are that clone and detection of the gene in cDNA use.
Pcr amplification product length is 744bp (Fig. 1), sequencing result such as SEQ in three kinds of materials in the gene cDNA
Shown in ID No.1；In gDNA the gene PCR amplified production length in purple and non-purple material be respectively 1665bp and
5442bp, gene sequencing result is as shown in SEQ ID No.5.
The ORF sequences of the gene are obtained using DNAMAN softwares and translate into amino acid sequence, such as SEQ ID No.3 institutes
Show.
During for detecting the gene, obtained amplified production is splined on 1.5% Ago-Gel and electrophoresis, through bromination
It is imaged after the dyeing of second ingot in gel imager, the results are shown in Figure 2, and the Chinese cabbage containing BrMYB2 genes amplifies one
The band of 1665bp, and the Chinese cabbage containing Brmyb2 genes then amplifies the band of a 5442bp.
Embodiment 4：Chinese cabbage Purpled traits codominant marker and the detection application of inclined maternal instinct mark
Large fragment insertion in the gDNA sequences of the acquired gene, the conserved sequence at insertion both ends are designed
The codominant marker 2 of heterozygosis strain can be detected to (Marker1 and Marker2), and the conserved sequence design among insertion
The inclined maternal instinct mark for going out the character that can combine purple Chinese cabbage segregating population is a pair of (Marker8).With what is extracted in embodiment 2
STb gene is masterplate, carries out the detection of Chinese cabbage Purpled traits codominant marker and inclined maternal instinct mark.
1) 20 μ l PCR reaction systems are：The template DNA of 50ng/ μ l is 2 μ l, 2 × Taq Master Mix 10 μ l, 10 μ
Each 1 μ l of upstream and downstream primer of M, reaction system is supplemented to 20 μ l with aseptic deionized water.
Primer sequence is：
Marker1-F：5’-TGGTGTACTTTGATCCTTCGTG-3’
Marker1-R：5’-ACTGCATTCGCGTCTCCTAC-3’
Marker2-F：5’-CTTTTCTGCACGAACCCG-3’
Marker2-R：5’-ATTCGCGTCTCCTACTCCATAT-3’
Marker8-F：5’-AAAAGGTGCATGGACTGCT-3’
Marker8-R：5’-TGTTTTGGTGTCGATGAAGAAG-3’.
2) pcr amplification reaction carries out in PCR instrument
Pcr amplification reaction condition is：94 DEG C of pre-degeneration 3min；Then 94 DEG C of denaturation 30s, 58 DEG C of annealing 30s, 72 DEG C are prolonged
Stretch, totally 35 circulations；Last 72 DEG C re-extend 8min, 4 DEG C of preservations.Wherein 72 DEG C extension 4min exist for Marker1 and Marker2
Detection in gDNA uses, and 72 DEG C of extension 1min detect use for Marker8 in gDNA.
During amplified production for detecting codominant marker, it is splined on 1.5% Ago-Gel and electrophoresis, through bromination second
It is imaged after ingot dyeing in gel imager.The results are shown in Figure 3 by codominant marker, the Chinese cabbage amplification containing BrMYB2 genes
Go out the band (Marker1 306bp, Marker2 348bp) of a treaty 300bp, and the Chinese cabbage containing Brmyb2 genes is then
Amplify the band (Marker1 4081bp, Marker2 4123bp) of an about 4000bp.And heterozygosis strain two couple mark
Note can amplify the band of two kinds of sizes.
During amplified production for detecting inclined maternal instinct mark, it is splined on 1.5% Ago-Gel and electrophoresis, through bromination second
It is imaged after ingot dyeing in gel imager.The results are shown in Figure 4, and the Chinese cabbage containing Brmyb2 genes then amplifies one
The band of 972bp, and the Chinese cabbage containing BrMYB2 genes is without special amplified production.If with the big of purple leaf-head character
Chinese cabbage can also amplify the band, then it is heterozygosis strain to illustrate the plant.
Embodiment 5：Chinese cabbage Purpled traits codominant marker and inclined maternal instinct mark the application in assisted Selection
It is right using codominant marker 2 using the F2 segregating populations of drinamyl Chinese cabbage and common Chinese cabbage crossing as object
(Marker1 and Marker2) and inclined maternal instinct mark a pair of (Marker8) carry out drinamyl Chinese cabbage assisted Selection.
1) 20 μ l PCR reaction systems are：The template DNA of 50ng/ μ l is 2 μ l, 2 × Taq Master Mix 10 μ l, 10 μ
Each 1 μ l of upstream and downstream primer of M, reaction system is supplemented to 20 μ l with aseptic deionized water.
Primer sequence is：
Marker1-F：5’-TGGTGTACTTTGATCCTTCGTG-3’
Marker1-R：5’-ACTGCATTCGCGTCTCCTAC-3’
Marker2-F：5’-CTTTTCTGCACGAACCCG-3’
Marker2-R：5’-ATTCGCGTCTCCTACTCCATAT-3’
Marker8-F：5’-AAAAGGTGCATGGACTGCT-3’
Marker8-R：5’-TGTTTTGGTGTCGATGAAGAAG-3’.
2) pcr amplification reaction carries out in PCR instrument
Pcr amplification reaction condition is：94 DEG C of pre-degeneration 3min；Then 94 DEG C of denaturation 30s, 58 DEG C of annealing 30s, 72 DEG C are prolonged
Stretch, totally 35 circulations；Last 72 DEG C re-extend 8min, 4 DEG C of preservations.Wherein 72 DEG C extension 4min exist for Marker1 and Marker2
Detection in gDNA uses, and 72 DEG C of extension 1min detect use for Marker8 in gDNA.
Amplified production is splined on 1.5% Ago-Gel and electrophoresis, after ethidium bromide staining in gel imager
Imaging.
3) PCR amplification result
The mark result and ball color detected to 400 F2 segregating populations single plants is completely the same, illustrates to utilize Chinese cabbage purple
It is feasible, especially inclined maternal instinct mark Marker8 and phenotype that character codominant marker and inclined maternal instinct mark, which carry out assisted Selection,
With reference to homozygous, heterozygosis drinamyl single plant simple and quick can be distinguished.
Nucleotides sequence list e-file
<110>Xibei Univ. of Agricultural ＆ Forest Science ＆ Technology
<120>Anthocyanin synthesis control gene and its application
<141>
<160>
<210>1
<211>744
<212>DNA
<213>BrMYB2 gene orders
<220>
<223>
<400>1
ATGGAGGGTTCGTCCCAAGGGTTGAAAAAAGGTGCATGGACTGCTGAAGAAGATAATCTCTTGAGGCAATGCA
TTGATAAGTATGGAGAAGGGAAATGGCACCAAGTTCCTTTAAGAGCTGGTCTAAATCGGTGCAGGAAGAGTTGTAGA
CTAAGATGGTTGAACTATTTGAAGCCAAGTATCAAGAGAGGAAAACTCAACTCCGATGAAGTTGATCTTCTTATTCG
CCTTCATAAGCTTTTAGGAAACAGGTGGTCTTTAATTGCTGGTAGATTACCCGGTCGGACCGCCAATGACGTCAAAA
ATTACTGGAACACCCATTTGAGTAAGAAACATGAACCGGGTTGTAAGACCCAGATGAAAAAGAGAAACATTCCTTGC
TCTTATACCACACCAGCCCAAAAAATCGACGTTTTCAAACCTCGACCTCGATCCTTCACCGTTAACAGCGGCTGCAG
CCATAATAATGGCATGCCAGAAGCTGGCATTGTTCCTCTATGCCTTGGACACAACGATACTAATAATGTTTCTGAAA
ATATAATCACATGTAACAAAGATGATGATAAATCTGAGCTTGTTAGTCATTTAATGGATGGTCAGAATAGGTGGTGG
GAAAGTTTGCTAGATGAGAGCCAAGATCCAGCTGCGCTCTTTCCAGAAACTACAGCAATAAAAAAGGGCGCAACCTC
CGCGTTTGACGTTGAGCAACTTTGGAGCCTGTTGGATGGAGAAACTGGAACTTGA
<210>2
<211>744
<212>DNA
<213>Brmyb2 gene orders
<220>
<223>
<400>2
ATGGAGGGTTCGTCCCAAGGGTTGAAAAAAGGTGCATGGACTGCTGAAGAAGATAATCTCTTGAGGCAATGCA
TTGATAAGTATGGAGAAGGGAAATGGCACCAAGTTCCTTTAAGAGCTGGTCTAAATCGGTGCAGGAAGAGTTGTAGA
CTAAGATGGTTGAACTATTTGAAGCCAAGTATCAAGAGAGGAAAACTCAACTCCGATGAAGTTGATCTTCTTATGCG
CCTTCATAAGCTTTTAGGAAACAGGTGGTCTTTAATTGCTGGTAGATTACCCGGTCGGACCGCCAATGACGTCAAAA
ATTACTGGAACACCCATTTGAGTAAGAAACATGAACCGGGTTGTAAGACCCAGATGAAAAAGAGAAACATTCCTTGC
TCTTATACCACACCAGCCCAAAAAATCGACGTTTTCAAACCTCGACCTCGATCCTTCACCGTTAACAGCGGCTGCAG
CCATAATAATGGCATGCCAGAAGCTGACATTGTTCCTCTATGCCTTGGACACAACGATACTAATAATGTTTCTGAAA
ATATAATCACATGTAACAAAGATGATGATAAATCTGAGCTTGTTAGTCATTTAATGGATGGTCAGAATAGGTGGTGG
GAAAGTTTGCTAGATGAGAGCCAAGATCCAGCTGCGCTCTTTCCAGAAACTACAGCAATAAAAAAGGGCGCAACCTC
CGCGTTTGACGTTGAGCAACTTTGGAGCCTGTTGGATGGAGAAACTGGAACTTGA
<210>3
<211>247
<212>Amino acid
<213>Brmyb2 amino acid sequences
<220>
<223>
<400>3
MEGSSQGLKKGAWTAEEDNLLRQCIDKYGEGKWHQVPLRAGLNRCRKSCRLRWLNYLKPSIKRGKLNSDEVDL
LIRLHKLLGNRWSLIAGRLPGRTANDVKNYWNTHLSKKHEPGCKTQMKKRNIPCSYTTPAQKIDVFKPRPRSFTVNS
GCSHNNGMPEAGIVPLCLGHNDTNNVSENIITCNKDDDKSELVSHLMDGQNRWWESLLDESQDPAALFPETTAIKKG
ATSAFDVEQLWSLLDGETGT
<210>4
<211>247
<212>DNA
<213>The nucleotide sequence of sense primer CVA10-F2
<220>
<223>
<400>4
MEGSSQGLKKGAWTAEEDNLLRQCIDKYGEGKWHQVPLRAGLNRCRKSCRLRWLNYLKPSIKRGKLNSDEVDL
LMRLHKLLGNRWSLIAGRLPGRTANDVKNYWNTHLSKKHEPGCKTQMKKRNIPCSYTTPAQKIDVFKPRPRSFTVNS
GCSHNNGMPEADIVPLCLGHNDTNNVSENIITCNKDDDKSELVSHLMDGQNRWWESLLDESQDPAALFPETTAIKKG
ATSAFDVEQLWSLLDGETGT
<210>5
<211>1665
<212>DNA
<213>BrMYB2 molecular labelings
<220>
<223>
<400>5
ATGGAGGGTTCGTCCCAAGGGTTGAAAAAAGGTGCATGGACTGCTGAAGAAGATAATCTCTTGAGGCAATGCA
TTGATAAGTATGGAGAAGGGAAATGGCACCAAGTTCCTTTAAGAGCTGGTATGTCTTTTTTTTTGATAACATAAGAG
CTGGTATGCTACTTTTATTAATTTTCACACACACACACACACACATATAACTAATAAGTACGTATATTCTTTTTATT
TTTCAGTACATTTATTCTCTTTCTCTCTGTCTAGTATTAGGAAATTAATTAACACCGGGGTACACAATCATTGTTTT
TCTTTTCGTTTTAATGAAGGAATCATAGATTCATATGTTCTAATGTTTTTCATGAAAAAAAAAACATTTGCGTTCTT
CATGTTTAATTACAAAGCGAGAAAATGTCAACTCTCTTTATTGATTCGTCGTTTTTCTTTTTTTTTTTGAGAAAGAG
CTTTTTTGATTAGTGAACTTTTCTGCACGAACCCGTGTGTTTGTGTGGAATATGTTGTTTATTCTGGTGTACTTTGA
TCCTTCATGATAAAATTTTACTTCCTTTGTTATTAAATATAAGATATTTTGGTAGAAGCAAACATATTAAGAAAACT
ATTTTTTGTCTAGAAAATATCATTAAAACTATAAATTAATGGTGTTCAACCAATTACAAAATAGACTATTAAAATAT
GATTGGGTTCACAGTTTTTAATAAAGTAAAAGTTACCTAGAAAATTGAAAACATTTTATATATTGGATAATTAAAAC
ATCAAAATTCAATAAAACATCCTATTTTTAGGAACATATGGAGTAGGAGACGCGAATGCAGTTTTTGCTCGTTCTTT
TAATAATATTAAATGTCAATTATTGGTTTTGTAGGTCTAAATCGGTGCAGGAAGAGTTGTAGACTAAGATGGTTGAA
CTATTTGAAGCCAAGTATCAAGAGAGGAAAACTCAACTCCGATGAAGTTGATCTTCTTATTCGCCTTCATAAGCTTT
TAGGAAACAGGTTTACATTCCAGACACAAATTCAACTGTATTTCGTATCCTCATTCGGTCTAATCTAATCATTTGAT
TTGTTTTTTTTTTTGATAAAAATACTTAAATTTATTTCATATGTAAATGATCCATTACTAAGTCAAATATATCCCTA
ATTTTTCAAATGCATGCTTAGGTGGTCTTTAATTGCTGGTAGATTACCCGGTCGGACCGCCAATGACGTCAAAAATT
ACTGGAACACCCATTTGAGTAAGAAACATGAACCGGGTTGTAAGACCCAGATGAAAAAGAGAAACATTCCTTGCTCT
TATACCACACCAGCCCAAAAAATCGACGTTTTCAAACCTCGACCTCGATCCTTCACCGTTAACAGCGGCTGCAGCCA
TAATAATGGCATGCCAGAAGCTGGCATTGTTCCTCTATGCCTTGGACACAACGATACTAATAATGTTTCTGAAAATA
TAATCACATGTAACAAAGATGATGATAAATCTGAGCTTGTTAGTCATTTAATGGATGGTCAGAATAGGTGGTGGGAA
AGTTTGCTAGATGAGAGCCAAGATCCAGCTGCGCTCTTTCCAGAAACTACAGCAATAAAAAAGGGCGCAACCTCCGC
GTTTGACGTTGAGCAACTTTGGAGCCTGTTGGATGGAGAAACTGGAACTTGA
<210>6
<211>5442
<212>DNA
<213>Brmyb2 molecular labelings
<220>
<223>
<400>6
ATGGAGGGTTCGTCCCAAGGGTTGAAAAAAGGTGCATGGACTGCTGAAGAAGATAATCTCTTGAGGCAATGCA
TTGATAAGTATGGAGAAGGGAAATGGCACCAAGTTCCTTTAAGAGCTGGTATGTCTTTTTTTTTTGATAACATAAGA
GCTGGTATGCTACTTTTATTAATTTTCACACACACACACACACACACATATAACTAATAAGTACGTATATTCTTTTT
ATTTTTCAGTACATTTATTCTCTTTCTCTCTGTCTAGTATTAGGAAATTAATTAACACCGGGGTACACAATCATTGT
TTTTCTTTTCGTTTTAATGAAGGAATCATAGATTCATATGTTCTAATGTTTTTCATGAAAAAAAACATTTGCGTTCT
TCATGTTTAATTACAAAGCGAGAAAATGTCAACTCTCTTTATTGATTCGTCGTTTTTCTTTTTTTTTTTGAGAAAGA
GCTTTTTTGATTAGTGAACTTTTCTGCACGAACCCGTGTGTTTGTGTGGAATATGTTGTTTATTCTGGTGTACTTTG
ATCCTTCGTGATAAAATTTTACTTCCTTTGTTCTTAAATATAAGATATTTTGGTAGAAGCATACATATTAAGAAAAC
TATTTTTTGTTTAGAAAATATCATTAAAACTATAAATTAATGGTGTTCAACCAATTACAAAATAGACTATTAAAATA
TGATTGGGTTCACAGTTTTTAATAAAGTAAAAGTTACCTAGAAAATTGAAAACATTTTATATCTAATACTATAAAGA
AGACTATTCTCTTCTTCTCAGCTTGCCACGTCACTGAATCGAATTCTAACAATGCGACACGTGTTCGTGCTCATGTT
ATACTCTGCGTTTCATTAAAGCTTGATGTGGGCTGGTGTTTTCAATTACGGTTTTGATACATATTGGATCTCAATAA
TAATAGGCCCATTTCAAAATCATGAGAGATGCAAGCCGATTAGGGTTTCTGCTCTTCTTCTTCATCGACACCAAAAC
AGTTCCCAAACAAAAATGATGGGAGTTTCGAGGTATCTCTTTGTTGATCTTCGATGGCGTCGTTCTGGTTACTCTCC
TAATTAGATGAAACGCTCCAAGTTACAGATCCATAGCCTTTATGACCTTCTTAACTCCAAAGACCGAAATGAGGCAC
GATCTGACGTTGAATGCGAGTTTACTCTCTGTGAGGCGGTGTAATCTACACAATATAAAGGTTAGCATCCTTCCCTT
GAACATCATCCTCGCAAATTCTAATAAAACTTCAAGGTAGTAGCTTTTGTTTGATGTTTATCAAACTCTAGGTTTTG
TTTGGTTCTTTGTCTTTCTATTTCCTCTCCCCAAGATCGTTAGATAAACATTTCGGATCGCTTTTGATAATTTTTTT
TGTCTTATCTACAGTTGGCTCAACACAAGCCACATGGTGATGGAAGATCTTTGCAACAGTCGTTCACTGTGAAGCGG
GGATTTTCGGAAGGCTATGGCAAAGATTCTGTGTGAGGAGATGAGTTGAGATGAGGATGAGAGTTCTTACTTTTCTT
TGAAGAAATTGATTGACGGTTCAAGACAATTACTCCATTAGCGCCCGCGAACCTGTGGGAAGAGCTTTCGTAGTGTT
TGGTTTACTCGCCGGTGGCGTTGCGGAAGTTCAATTCGTCGGCACCGTTTAAATGGCAACACCGACTCACTTCATCT
TCATCTCTTTGGGACGACATCTGCGCCGTGGTAACATGCTTTCTTATTGTCGGTGTGGTGACGTCAGCCTCTCCACA
CATTCAAACGCATGTTATTGACAATCAAATTATTATGCAGGCTTTCTCTCAACATACAGAGCTTCCATAGTTATTGT
TAAATCTTAATCATGTCTGAGGATACCATCACCGTCCCTCTCAGCTGCTGGAATGTACTCAGCTTGGTTGGATGGAG
CGTCTGCTTGGCTGGAATTCAGAAAGTTCTGCTCCAAAGGAACCTGTCTGAACGTCAATGATGCCTTTGGCCAAGAA
TTATAACATGGTGGCTGATCCAAGAAATACCTGAACATTGCAATCTTCTTAAGAGCACCGAATGTAACATGTGACGA
GGTGTCAAAAGCTCTTCTAGATTGTAAGATTGGTCTCCCATTTGGTTTGTTACTTAGGGGTAAAGTTCTATTTGCTT
TGATCACTTCAACTTGCAAAATAGTGTTAACGGAACACATTCTATGCACTAATCTTATTTCCATCATTTTCTATTCA
CTCCACTAATCAAGTTCAGTCCAACGACTATTTATCATCCAGGAATCAAGTTCTATTTGTTAAATATATTTGTGAAT
CACACACAGCCAAGCTATCCACAAATTTATTGAACAATCTCTTCAAAGACAAACTTATAATACACTGCCACTTTCAA
AAAATGTTTTGTGTACACTTCTTTGGACAGACGAAAACTATGACCGTCTTATTAATGGTAAGTCTGGCTCAAACCAA
GCATCAATGCCATCACGTTCCTGCTCTTATTATTCTGAAAGGAGCAAGAGGAAAGCAGGTTGTAGCACAGTCATGTT
CCAGCTCCATATTTCACAGAAGCCATCATTGCCGAGTTGAGTTGCAGTATTGTGTAAAAGTTTCTAACCACGAGGAG
TTTAGTGTTGGAAAATATATTGGAGGTGCAGTGTTTATACCGAAAAGTTCTCAGGCAGAGATATTGTGTAGGTCTAA
ATGTCATAAAAGGACAGAAAATATAGTAGATGGAATTCTGTTTTTGTGTCCAGTTATATTCTCCGACTGTAGCTTGA
AGAGCAAACTATCTAAGAGAGGTGAGATTGGTTTTAATCTGCCATTTCTGGAGAGTTGTGAAGATGTTGAAGGAGAG
CAAGGTGAAGAATGTGATTGCTCACGCTGGGTTGTAAGAGACAGTCTTAGCTATTGGTTTCTGATTCTCAGATTTTT
CAGCTGCAAATCTCAGACCGGTTTCTAAACACCAAAACAGCCTTTGGAAGACTTTCTTTTGTTGTGTGTTCTCCTGG
CGGTGATGGGCCTACGGATCGCTGTGGTGACAGTGCAAAGGGGAGTGTTGTCGGGTGGTTATCAATTCAATACTATA
CAATTCTTTGCTTACAAAGCTGTGTAAGAGTATATTGTTATTATATAATGGTGAAAGAGTCGAAGTTGATGCTCACT
GGAAGGAAATGAAGTAATGGAAAAAAAAACTCCACTAGATGCAAGAAACAATAGTTTTCTAAGTAGCGTTGAAATAT
TATCAATAAAAATAAAAGTATATAACAAACAATGTTTATTTGATAAGATTTCTAAACAATAAAGAGTGTTAAAGATT
CGGCAAATAAATTATTTAGAATGTACATGATACAAAATATTGCAAAGTTTTTTTTTTTTTTTTGCTAAAAGTAAATG
TGTTGTGTTGATTTAAAATATTGCAAAGTTGTTATGAAAAATGTAATATCTGAAGATACCATCGAAAACATTAAAAC
ATTAAATTTTAACACATTTCCAACAGTTAAAAATATCAATTATCTTATATTATAAATGAATATGCTACGCAAAAAAT
GTACAATAATTCTTTTTATCAGCTACAGAAAACAGTTATGAAATTTTGTAACAAACATCAAATGTATTATTAACCGA
AAGTCAACATTTTATCTATTAAAAAGTATTATACATGCTTATCAACGTCCCTACCACAAATTATAACTAATTAAATC
AAGTTGAAGCCAATATCTAATACTCAATATTCTTTGTTCCAAAATCTTAATAATAAAAATGAAAAAATATTAGCAAA
ACATTGTTTACTTTTAATTTATTTTTGAAAATTATTTAGTATTAGTTAAGTTTATTAAATATTTAATTTGATATTGT
AAATTTGAATATATACAATCAAATACACATTTTTAAAAAAATAGATGAAAGTATTTTTAAAACACGTATGTGTACCT
AATAAATATGCAAAAAATCATACATGATAACTTTTTAAAAATAAAAAGCAATTAAATGTTATATGTAGATTTATTAT
TTATTATTTATGTCATTCAGATAAAAATATAGCAAAGGTGTAATAATATTCATAATATATACTAGAAAATTATATGC
ATCATTAAATTATATATTACTAAAAACTAATTTAAATTTTATAAACAATAAAATATAATTATGAAAGTAGTTTATAT
AATGAATAAAATATACAAAATTATAGTTAATTTAAATTAATAATGATATGAAAATAAAAATATGTCTTTACATAATC
GAACAAAGAAAAAAATATAAAATAGTCTGACAACAAACAAAGATAGAAGAAATGAAATTAGTTGATTTATACGAAAC
AAATAAGTTAAAATATATATACTAACATAATAAAAATATAATACAATATTATTAAAAATCACAATAATTGTATTAGA
AACCTACTAAATAGTAACAAAACTGACTTGAAAGAAAACAATTTTAAAAATAATGACCTGGATAAAATAATCATTTT
AAAACAAAAATTTTAAAAATTATTTAATTTTCCGCCCGCCCGTAGGGCGGGTTTACTCTAGTATTGGATAATTGAAA
CATCAAAATTTTCAGTAAAACATCCTATTTTTAGGAACATATGGAGTAGGAGACGCGAATGCAGTTTTTGCTCGTTC
TTTTAATAATATTAAATGTCAATTATTGGTTTTGTAGGTCTAAATCGGTGCAGGAAGAGTTGTAGACTAAGATGGTT
GAACTATTTGAAGCCAAGTATCAAGAGAGGAAAACTCAACTCCGATGAAGTTGATCTTCTTATGCGCCTTCATAAGC
TTTTAGGAAACAGGTTTACATTCAAGACACAAATTCAACTGTATTTCGTATCCTCATTCGGTCTAATCTAATCATTT
GATTTGTTTTTTTTTTTGATAAAAAGTACTTAAATTTATTTCATATGTAAATGATCCATTACTAAGTCAAATATATC
CCTAATTTTTCAAATGCATGCTTAGGTGGTCTTTAATTGCTGGTAGATTACCCGGTCGGACCGCCAATGACGTCAAA
AATTACTGGAACACCCATTTGAGTAAGAAACATGAACCGGGTTGTAAGACCCAGATGAAAAAGAGAAACATTCCTTG
CTCTTATACCACACCAGCCCAAAAAATCGACGTTTTCAAACCTCGACCTCGATCCTTCACCGTTAACAGCGGCTGCA
GCCATAATAATGGCATGCCAGAAGCTGACATTGTTCCTCTATGCCTTGGACACAACGATACTAATAATGTTTCTGAA
AATATAATCACATGTAACAAAGATGATGATAAATCTGAGCTTGTTAGTCATTTAATGGATGGTCAGAATAGGTGGTG
GGAAAGTTTGCTAGATGAGAGCCAAGATCCAGCTGCGCTCTTTCCAGAAACTACAGCAATAAAAAAGGGCGCAACCT
CCGCGTTTGACGTTGAGCAACTTTGGAGCCTGTTGGATGGAGAAACTGGAACTTGA
<210>7
<211>19
<212>Artificial sequence
<213>Sense primer
<220>
<223>
<400>7
5’- ATGGAGGGTTCGTCCCAAG-3’
<210>8
<211>22
<212>Artificial sequence
<213>Anti-sense primer
<220>
<223>
<400>8
5’-TGGTGTACTTTGATCCTTCGTG-3’
<210>9
<211>22
<212>Artificial sequence
<213>Marker1-F
<220>
<223>
<400>9
5’-TGGTGTACTTTGATCCTTCGTG-3’
<210>10
<211>19
<212>Artificial sequence
<213>Marker1-R
<220>
<223>
<400>10
5’-ACTGCATTCGCGTCTCCTAC-3’
<210>11
<211>18
<212>Artificial sequence
<213>Marker2-F
<220>
<223>
<400>11
5’-CTTTTCTGCACGAACCCG-3’
<210>12
<211>22
<212>Artificial sequence
<213>Marker2-R
<220>
<223>
<400>12
5’-ATTCGCGTCTCCTACTCCATAT-3’
<210>13
<211>19
<212>Artificial sequence
<213>Marker8-F
<220>
<223>
<400>13
5’-AAAAGGTGCATGGACTGCT-3’
<210>14
<211>22
<212>Artificial sequence
<213>Marker8-R
<220>
<223>
<400>14
5’-TGTTTTGGTGTCGATGAAGAAG-3’
Claims (10)
1. Anthocyanin synthesis control gene, it is characterised in that the sequence of the gene is：
(1)SEQ ID NO:Nucleotide sequence shown in 1；Or
(2)SEQ ID NO:Nucleotide sequence shown in 1 is substituted, lacks and/or increases one or more nucleotide and expression
The nucleotide sequence of identical function protein；Or
(3) there is more than 90% homology with the nucleotide sequence of (1) or (2) and expresses the nucleotides sequence of identical function protein
Row.
2. expression cassette, expression vector, cloning vector or engineering bacteria, it is included comprising the nucleic acid described in claim 1.
3. gene described in claim 1 synthesizes the application in anthocyanidin in plant, the plant includes：Chinese cabbage, not balling
Chinese cabbage, arabidopsis.
4. a kind of construction method of genetically modified plants, it is characterised in that using genetic engineering means, be overexpressed in target plant
Gene described in claim 1, screens positive transgenic plant.
5. with the relevant molecular labeling of Chinese cabbage anthocyanidin synthetic gene, it is characterised in that include one in the molecular labeling
The insertion mutation of 3772bp fragments, its gDNA sequence are as shown in SEQ ID No.5.
6. the PCR detection kit for identifying drinamyl Chinese cabbage material, it is characterised in that the kit, which contains, to be useful for expanding
The sense primer and anti-sense primer of molecular labeling described in claim 5：
Sense primer：5’-ATGGAGGGTTCGTCCCAAG-3’
Anti-sense primer：5’-TCAAGTTCCAGTTTCTCCATCC-3’.
7. the PCR detection kit for identifying drinamyl Chinese cabbage material, it is characterised in that the kit, which contains, to be useful for expanding
The sense primer and anti-sense primer of molecular labeling described in claim 5：
Marker1-F：5’-TGGTGTACTTTGATCCTTCGTG-3’
Marker1-R：5’-ACTGCATTCGCGTCTCCTAC-3’
Marker2-F：5’-CTTTTCTGCACGAACCCG-3’
Marker2-R：5’-ATTCGCGTCTCCTACTCCATAT-3’
Marker8-F：5’-AAAAGGTGCATGGACTGCT-3’
Marker8-R：5’-TGTTTTGGTGTCGATGAAGAAG-3’.
8. application of the molecular labeling described in claim 5 in purple Chinese cabbage molecular mark.
9. application of the molecular labeling described in claim 5 in drinamyl Chinese cabbage material is identified.
10. application as claimed in claim 9, it is characterised in that comprise the following steps：
(1) DNA of Chinese cabbage to be measured is extracted；
(2) using DNA as template, pcr amplification reaction is carried out using sense primer and anti-sense primer；
(3) pcr amplification product is detected.
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| CN201711269495.8A CN108018290B (en) | 2017-12-05 | 2017-12-05 | Plant anthocyanin synthesis control gene and application thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113439657A (en) * | 2020-03-24 | 2021-09-28 | 西北农林科技大学 | Breeding method of new germplasm of purple-orange Chinese cabbage |
| WO2021189221A1 (en) * | 2020-03-24 | 2021-09-30 | 西北农林科技大学 | Selective breeding method for new germplasm of purple-orange chinese cabbage |
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|---|---|---|---|---|
| WO2003100068A1 (en) * | 2002-05-29 | 2003-12-04 | Aresa Biodetection Aps | Reporter system for plants |
| CN102120762A (en) * | 2010-05-14 | 2011-07-13 | 昆明理工大学 | Arabidopsis anthocyanin pap1 purified protein and preparation method and applications thereof |
| CN104531754A (en) * | 2014-12-19 | 2015-04-22 | 西南大学 | Application of gene for interfering expression of LCYB and LCYE and simultaneously over-expressing PAP in preparation of brassica plant having red petals |
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2017
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| WO2003100068A1 (en) * | 2002-05-29 | 2003-12-04 | Aresa Biodetection Aps | Reporter system for plants |
| CN102120762A (en) * | 2010-05-14 | 2011-07-13 | 昆明理工大学 | Arabidopsis anthocyanin pap1 purified protein and preparation method and applications thereof |
| CN104531754A (en) * | 2014-12-19 | 2015-04-22 | 西南大学 | Application of gene for interfering expression of LCYB and LCYE and simultaneously over-expressing PAP in preparation of brassica plant having red petals |
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| NING GUO等: "Anthocyanin biosynthetic genes in Brassica rapa", 《BMC GENOMICS》 * |
| 匿名: "PREDICTED: Brassica rapa transcription factor MYB114 (PAP1), transcript variant X1, mRNA, Accsession ID:XM_009106954.2", 《GENBANK数据库》 * |
| 张惠展: "《基因工程（第4版）》", 31 January 2017 * |
| 李益等: "紫色芸薹属蔬菜花青素合成调控研究进展", 《山东农业科学》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113439657A (en) * | 2020-03-24 | 2021-09-28 | 西北农林科技大学 | Breeding method of new germplasm of purple-orange Chinese cabbage |
| WO2021189221A1 (en) * | 2020-03-24 | 2021-09-30 | 西北农林科技大学 | Selective breeding method for new germplasm of purple-orange chinese cabbage |
| US20220304265A1 (en) * | 2020-03-24 | 2022-09-29 | Northwest A&F University | A method for breeding new purple-orange chinese cabbage germplasm |
| CN113439657B (en) * | 2020-03-24 | 2022-12-02 | 西北农林科技大学 | Breeding method of purple-orange Chinese cabbage germplasm |
Also Published As
| Publication number | Publication date |
|---|---|
| CN108018290B (en) | 2020-11-10 |
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