CN102690811B - Molecular marker SIsv1363 closely linked with millet leaf color gene - Google Patents
Molecular marker SIsv1363 closely linked with millet leaf color gene Download PDFInfo
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Abstract
The invention belongs to biology field, relate to a kind of molecular marker, concrete, relate to a kind of and the millet closely linked molecular marker of leaf color gene, does this molecular marker contain Seq? ID? sequence shown in No.1. The invention still further relates to the amplification primer of this molecular marker, this molecular marker and primer purposes in millet leaf color gene mapping or millet genetic breeding and a kind of Millet Breeding method. Genomic dna sequence and millet leaf color genetic link are got up by the molecular marker of the present invention, are conducive to the foundation of millet molecular mark system; The hereditary close linkage distance of described molecular marker and millet leaf color gene is 0.15cM. The molecular marker of the present invention and molecular marker amplimer can be applied to Millet Breeding practice and resource and cultivar identification easy, quickly, high flux.
Description
Technical field
The invention belongs to biology field, relate to a kind of molecular marker, particularly relate to a kind of and the millet closely linked molecular marker of leaf color gene. The invention still further relates to the amplification primer of this molecular marker, this molecular marker and primer purposes in millet leaf color gene mapping or millet genetic breeding.
Background technology
China is the country of origin of millet (SetariaitalicaL.Beauv.), is that the concentration of millet in the world plants state, and millet occupies an important position in the national economy and social production of China, and dry farming ecological agriculture construction is significant. Therefore, the breeding process accelerating millet is particularly important. Owing to millet is region importance crop, therefore current relatively backward about research means and the method for millet, how the research means of application of advanced science does Millet Breeding well is a severe problem. Along with molecular biological development, the appearance of molecular marking technique, genetic research and breeding for millet open new thinking and means. Exploitation has the molecular marker of the important character gene of China's characteristic and carries out assisted selection research, will play facilitation to improving China's Millet Breeding level.
Leaves of plants color is the general performance of various pigments in chloroplast, and normal blade Determination of Chlorophyll is preponderated, and is usually expressed as green. Leaf variegation be in higher plant mutation frequency higher and be prone to identify the mutant character. Up to now, in nearly all higher plants such as Oryza sativa L., Semen Tritici aestivi, Fructus Hordei Vulgaris, corn and soybean, Cotton Gossypii, Nicotiana tabacum L., Semen Maydis, Helianthi, Fructus Lycopersici esculenti, Fructus Cucumidis sativi, Rhizoma Solani tuber osi, mulberry, leaf color mutant has been had been found that. Its mutant gene directly or indirectly affects photosynthetic pigments, particularly chlorophyllous synthesis and degraded, causes that the content of various pigment and ratio change, thus causing leaves of plants chromatic variation. In recent years, the value of leaf color mutant increasingly receives publicity, now become the special material of research photosynthesis of plant mechanism, the growth of chlcrophyll biosynthesis approach, chloroplast and genetic control mechanism, while leaf color mutant also there is important using value in the utilization of the breeding of high photosynthetic efficiency and mark property. Some leaf variegations are taken as mark property and produce for stock breeding and cenospecies. Along with completing that arabidopsis, Oryza sativa L. isotype Plant Genome check order, leaf color mutant researching value in functional genomics receives more and more attention, and they have become the ideal material of research photosynthetical system structure, gene function and regulatory mechanism thereof. According to incompletely statistics, the related gene of the Oryza sativa L. Leaf color mutant of report alreadys more than 80 at present, and these mutants are broadly divided into the 5 big classes such as albinism, yellow type, stripe, green changing type, zebra leaf. But the research for millet Ye Sejiyin and closely linked molecular marker with it is substantially free of bibliographical information.
Summary of the invention
It is an object of the invention to provide a kind of and the millet closely linked molecular marker of leaf color gene.
It is a further object of the present invention to provide a kind of primer pair that can be used for pcr amplification and the millet closely linked molecular marker of leaf color gene and the molecular marker obtained by this primer pair amplifies.
Another object of the present invention is to provide the detection method of above-mentioned molecular marker purposes in millet leaf color gene mapping, detection and millet assistant breeding and above-mentioned molecular marker.
The purpose of the present invention also includes providing a kind of recombinant vector including above-mentioned molecular marker and the reconstitution cell containing this recombinant vector; The localization method of a kind of millet leaf color gene including using above-mentioned molecular marker is provided, and uses the millet auxiliary breeding means of described molecular marker.
To achieve these goals, present invention employs techniques below scheme:
The invention discloses a kind of and the millet closely linked molecular marker of leaf color gene, described molecular marker contains sequence shown in SeqIDNo.1; Preferred described molecular marker has sequence shown in SeqIDNo.1.
The invention also discloses the primer pair of a kind of amplification and the millet closely linked molecular marker of leaf color gene, the primer 1 of described primer pair is containing sequence shown in SeqIDNo.2, and primer 2 contains sequence shown in SeqIDNo.3; Preferred described primer 1 has sequence shown in SeqIDNo.2, and primer 2 has sequence shown in SeqIDNo.3;
SeqIDNo.2:5 '-GACATTCCAGTCGAGAAGCT-3 ';
SeqIDNo.3:5 '-GGTAGCTTCACTTCACAGAC-3 '.
The invention also discloses a kind of and the millet closely linked molecular marker of leaf color gene, described molecular marker is to be obtained for template through pcr amplification with the millet genomic DNA that blade is partially green by above-mentioned primer pair.
The preferred molecular marker obtained by above-mentioned primer pair amplifies contains sequence shown in SeqIDNo.1.
In one embodiment of the invention, described molecular marker (DNA fragmentation containing nucleotide sequence shown in SeqIDNo.1) is the DNA fragmentation of nucleotide sequence shown in SeqIDNo.1 in millet genome, nucleotide sequence beyond the 5 ' ends of the SeqIDNo.1 namely comprised and/or 3 ' ends is also the sequence in millet genome, preferably, for 5 ' end and/or 3 ' the upstream and downstream sequences held of SeqIDNo.1 in millet genome. As long as this molecular marker that it will be understood by those skilled in the art that in amplification or the partially green millet genomic DNA of detection blade, necessarily can detect or expand and obtain containing the sequence shown in SeqIDNo.1. 5 ' the ends of SeqIDNo.1 and/or the length of 3 ' the upstream and downstream sequences held are suitable length, it is not particularly limited, such as, meet the length of molecular marker less than 10,000bp, less than 5,000bp, less than 2,000bp, less than 1,500bp, less than 1,200bp, less than 1,000bp or less than 800bp.
In one embodiment of the invention, 5 ' the ends of the SeqIDNo.1 that described molecular marker (DNA fragmentation containing nucleotide sequence shown in SeqIDNo.1) comprises and/or 3 ' are held and have been operably connected artificial sequence and/or have controlled sequence, for instance promoter, enhancer, terminator, restriction enzyme site, primer sequence etc. Wherein, term " operationally " is defined as a kind of following conformation in the present invention, in this conformation, controls sequence such as promoter and is appropriately placed on a position of SeqIDNo.1, so that the generation of the polypeptide of the sequence-directed SeqIDNo.1 coding of this control.
The invention also discloses a kind of recombinant vector, it contains the molecular marker of the present invention. Described recombinant vector can be expression vector or the cloning vehicle of the molecular marker being inserted with the present invention. After obtaining above-mentioned recombinant vector, it will be understood by those skilled in the art that according to different needs, recombinant vector is transformed in suitable cell, obtain the reconstitution cell containing this recombinant vector. Therefore, the invention also discloses a kind of reconstitution cell containing described recombinant vector.
The preparation method that the invention also discloses the molecular marker of the present invention, comprise the steps: that the genomic DNA of the millet using leaf color partially green is as template, carrying out pcr amplification with above-mentioned primer pair, namely the 731bp amplified production obtained contains described molecular marker; Preferably, the step being purified by pcr amplification product is also included.
To those skilled in the art, it is possible to understand that, it is also possible to the method for DNA chemosynthesis obtains the molecular marker of the present invention.
The invention also discloses the detection method of described molecular marker, including step: design primer according to the nucleotide sequence of above-mentioned molecular marker, expand for template with detected millet genomic DNA, and judge whether amplified production exists this molecular marker. Preferably, described primer is the above-mentioned primer pair containing SeqIDNo.2 and SeqIDNo.3 respectively.
For example, it is possible to the genomic DNA of detected millet for template, carry out pcr amplification with above-mentioned primer (SeqIDNo.2 and SeqIDNo.3), obtain amplified production. Can be undertaken checking order or gel electrophoresis by the amplified production obtained.
The invention also discloses described molecular marker purposes in millet leaf color gene mapping or detection.
A kind of method that the invention also discloses millet leaf color gene mapping, described method includes the step using the molecular marker of the present invention.
The invention also discloses described molecular marker purposes in millet assistant breeding.
The invention also discloses a kind of millet auxiliary breeding means, described method includes the molecular marker of the detection present invention or the step of molecular marker primer pair.
The molecular marker of the present invention can be used in molecular mark from now on, it will be appreciated by those skilled in the art that, as by detect whether the molecular marker that there is the present invention to screen millet blade whether contain control leaf color be partially green color gene (such as, it is referred to, DNA molecular marker purposes in wheat breeding for disease resistance, Long Dong institute journal (natural science edition), volume the 1st phase April the 16th in 2006, P65-69). Described detection can be the method for PCR detection, specifically, it is possible to use the primer pair of the molecular marker of the above-mentioned present invention. Described detection can also be undertaken by sequence measurement. This millet auxiliary breeding means has simplicity, advantage quick, high-throughout.
In the present invention, specifically, described millet can be the F2 generation that a paddy No. 1, millet A2 sterile line, a confused flour beetle 3 or No. 3 selfings of a confused flour beetle produce. Wherein, the part F2 opening paddy No. 1, confused flour beetle 3 or No. 3 selfings generations of a confused flour beetle is partially green for leaf color; The part F2 that millet A2 sterile line, No. 3 selfings of a confused flour beetle produce is for the inclined yellow of leaf color.
Owing to adopting above technical scheme, make what the present invention possessed to have the beneficial effects that:
The invention provides and the millet closely linked molecular marker of leaf color gene, genomic dna sequence and millet leaf color genetic link are got up by this molecular marker, are conducive to the foundation of millet molecular mark system; The hereditary close linkage distance of described molecular marker and millet leaf color gene is 0.15cM. The molecular marker of the present invention and molecular marker amplimer can be applied to Millet Breeding practice and resource and cultivar identification easy, quickly, high flux.
Accompanying drawing explanation
Fig. 1: molecular marker primer (SeqIDNo.2 and the SeqIDNo.3) partial results to 480 the individual plant amplifications of F2 generation. Wherein:
Swimming lane 1-12 is the pcr amplification product of the individual plant that 12 strain millet leaf colors are partially green in F2 generation; Swimming lane 13-24 is the pcr amplification product of the individual plant of the 12 inclined yellow of strain leaf color in F2 generation. Swimming lane M is marker, and it is 100bpDNALadder; Its molecular weight includes:, 1500bp, 1000bp, 900bp, 800bp, 700bp, 600bp, 500bp, 400bp, 300bp, 200bp and 100bp.
Detailed description of the invention
The invention discloses a kind of primer pair and molecular marker SIsv1363 closely linked with millet leaf color gene. Utilize the primer pair of the present invention, carry out PCR with millet genomic DNA for template, it is possible to obtaining and the millet closely linked molecular marker of leaf color gene, this molecular marker is called after molecular marker SIsv1363 in the present invention. It is pointed out that except the molecular marker that it will be understood by those skilled in the art that except being obtained the present invention by above-mentioned pcr amplification, it is also possible to obtained the molecular marker of the present invention by chemosynthesis.
The primer pair of the present invention respectively containing sequence shown in ordered list SeqIDNo.2 and SeqIDNo.3,
SeqIDNo.2:5 ' GACATTCCAGTCGAGAAGCT-3 ';
SeqIDNo.3:5 '-GGTAGCTTCACTTCACAGAC-3 '.
Those skilled in the art know, in sequence shown in above-mentioned SeqIDNo.2 and SeqIDNo.3,1��10 base can be increased respectively at its 5 ' end or 3 ' ends, the base type increased can be determined according to the base type in the region that matches with SeqIDNo.2 and SeqIDNo.3 on millet genomic DNA foundation basepairing rule, the primer pair thus obtained essentially identical with the amplified production of SeqIDNo.2 and SeqIDNo.3 (DNA sequence between upstream and downstream primer is identical). Therefore, the above-mentioned 5 ' ends at SeqIDNo.2 and SeqIDNo.3 or 3 ' ends increase by 1��10 base respectively and can expand the primer pair obtaining essentially identical DNA fragmentation, are included in the primer pair of the present invention. In specific embodiment of the present invention, the primer pair of the present invention is preferably sequence shown in SeqIDNo.2 and SeqIDNo.3. The present invention is by the homozygote hybridization of partially green and maternal for the male parent blade inclined yellow of blade, it is thus achieved that F1 generation (confused flour beetle 3), then produces F2 for colony with F1 generation selfing, totally 480 individual plants. Preferred employing SV molecular markers development method, first carries out denovo order-checking (60XcontigN50:22K, scaffoldN50:320K to male parent; Totalsize:400Mb), female parent is resurveyed sequence (10X); Then according to Parent sequencing data, utilize the sequence difference between the SOAP comparison Parent of Hua Da independent development, respectively in 5 ' ends of male parent diversity sequence and 3 ' about 50bp positions, end outside, randomly select the primer of the Design of length diversity sequence amplification of about 20bp, devise 1105 pairs of primers according to different diversity sequences. Expand for template with the DNA of Parent and F1, from 1105 pairs of primers, filter out 616 pairs of primers with polymorphism and effectiveness. Adopt the 616 pairs of primers developed, 480 individualities of F2 colony are carried out PCR detection, and carries out data statistic analysis, carry out genetic map drafting with MapMaker3.0 software, obtain molecular marker closely linked with leaf color gene of the present invention and amplimer thereof. Adopt the male parent sequence that the primer pair amplifies filtered out obtains, i.e. molecular marker SIsv1363 in the present invention. F2 individuality is carried out character analysis, and according to gene character data and phenotype trait data by millet leaf color gene mapping on genetic map.
Below by specific embodiment and in conjunction with accompanying drawing, the present invention is described in further detail. The present invention is only further detailed by following example, should not be construed as limitation of the present invention.
Embodiment 1: millet F2 is for the structure of segregating population
Male parent: anti-Sethoxydin, plant type is high, and boot leaf is long and narrow, and bristle is red, and grain husk shell is red, can educate, and leaf colour cast is green, pollen yellow-white, and heading stage is late period. Male parent is No. 1 seed of paddy.
Maternal: not anti-Sethoxydin, plant type is short, boot leaf short-wide, and bristle is green, and grain husk shell is green, partial sterility, and leaf colour cast is yellow, and pollen is brown, and heading stage is early stage. Female parent is millet A2 male-sterile seed.
F2 informative population: male parent and hybridization of female parent obtain F1 generation (F1 leaf color is for partially green), and F1 selfing obtains F2. Wherein F1 is No. 3 seeds of a confused flour beetle. There are F2 for individual plant 480 strain, No. 1 seed of above-mentioned paddy, millet A2 male-sterile seed and No. 3 seed of confused flour beetle can referring to Chinese patent application " molecular marker SIsv0372s closely linked with millet anti-herbicide gene ", publication number CN101974521A, date of publication on February 16th, 2011.
Embodiment 2: Parent and F1 generation, F2 are for the extraction of genes of individuals group DNA
Extract the genomic DNA of the Parent in embodiment 1, F1 generation and 480 F2 generation individualities respectively by CTAB method, concrete grammar is as follows:
(1) weigh the fresh blade of 1.0g, shred and put into mortar, with adding 3mL1.5 �� CTAB after liquid nitrogen grinding, grind to form in the centrifuge tube that homogenate proceeds to 15mL, in mortar, then add 1mL1.5 �� CTAB flushing proceed to again in centrifuge tube. In 65 DEG C of water-bath 30min after mixing, period slowly shakes up frequently.
Wherein 1.5 �� CTAB formula following (1L):
Add deionized water and be settled to 1L, before using, add the mercaptoethanol of final concentration of 0.2% (2ml).
(2) it is cooled to room temperature, adds equal-volume chloroform/isoamyl alcohol (24: 1), mix gently, become bottle green to subnatant.
(3) the centrifugal 10min of 4200rpm, moves on to upper water new 15mL centrifuge tube mutually, adds the dehydrated alcohol of 2 times of volume pre-coolings, mix static 5min. Place 30min in-20 DEG C and precipitate DNA.
(4) the centrifugal 10min of 4200rpm, discards supernatant, adds 1mL75% washing with alcohol and precipitates 1 time, is inverted centrifuge tube dry DNA, adds 200 �� LTE dissolving DNAs.
(5) genomic DNA is detected with the agarose gel of 0.8%.
(6) the genomic DNA individual Parent obtained and F1 generation, F2 generation is stored in-20 DEG C standby.
Embodiment 3: the preparation of molecular marker
(SeqIDNo.2 and SeqIDNo.3), for template, is carried out pcr amplification with molecular marker amplimer by the male parent, F1 generation or the genomic DNA in F2 generation that extract in embodiment 2.
PCR reaction system is as follows:
PCR response procedures is as follows:
94 DEG C of denaturations 5 minutes; 94 DEG C of degeneration 30 seconds, anneal 30 seconds for 60 DEG C, and 72 DEG C extend 40 seconds, run 35 circulations; Last 72 DEG C extend 3 minutes. Pcr amplification product can 4 DEG C of preservations.
Molecular marker is obtained, it is preferable that after amplification, amplified production is purified operation through above-mentioned amplification procedure. Checking order after purification, result is such as shown in SeqIDNo.1.
To those skilled in the art, it is possible to understand that, it is also possible to obtain this molecular marker by the method for DNA chemosynthesis.
Embodiment 4:SV molecular markers development
Male parent: denovo checks order, 60XcontigN50:22K, scaffoldN50:320K; Totalsize:400Mb; Maternal: resurvey sequence 10X.
According to Parent sequencing data, utilize SOAP software (the such as SOAP2.20 of Hua Da independent development, can download from http://soap.genomics.org.cn/, other sequence alignment program can also be used) sequence difference that compares between Parent, it is then based on the sequence of difference, expands the primer of diversity sequence by primerpremier software design; Based on different diversity sequences, devise 1105 pairs of primers altogether. Table 1 below has illustrated part primer sequence (SeqIDNo.2-SeqIDNo.41) therein
Table 11105 is to the part primer in random primer
Respectively with the genomic DNA of the Parent extracted and F1 generation for template, carry out pcr amplification with 1105 pairs of primers of design.
PCR reaction system (25 �� L):
PCR response procedures: 94 DEG C of denaturation 5min; Subsequently into 35 circulations: 94 DEG C of degeneration 30s, 60 DEG C of annealing 30s, 72 DEG C extend 40s; After loop ends, 72 DEG C extend 3min; 4 DEG C of preservations.
PCR primer electrophoresis detection: agarose gel 120v electrophoresis 25min, the EB dyeing 10min of 1.2%, according to glue record.
The effectiveness of primer and polymorphism: refer to whether there is amplified production at this effectiveness, polymorphism refers to that between Parent, the clip size of amplified production is variant.
The screening of primer is carried out: Parent and F1 all have amplified production according to following screening criteria, and Parent amplified production all only has a distinct banding pattern and size variant, F1 shows as the heterozygosis banding pattern of Parent banding pattern, namely has male parent and two bands of maternal banding pattern.
The selection result: according to above-mentioned screening criteria, filter out 616 pairs of primers from 1105 pairs of primers of design.
Embodiment 5: genetic map construction and gene mapping
(1) genetic map construction
With 616 couple of exploitation, there is the molecular marker of polymorphism and 480 individualities of F2 colony are carried out PCR detection, 480 individual genomic DNAs of the template used F2 colony for preparing.
PCR primer is carried out agarose gel electrophoresis, obtains the result of the individual amplification of molecular marker primer pair 480.
Whole electrophoresis result are carried out data statistic analysis, concrete grammar is as follows: by F2 colony individual plant amplified band be male parent type be designated as a, amplified band be maternal type be designated as b, what amplified band contained male parent type and maternal type simultaneously is designated as h, band Fuzzy or disappearance be designated as-, be equivalent to shortage of data, finally give the genotype data of 480 individual 616 pair primer amplifications of F2 colony. Such as, 480 the individual data obtained with pair of primers are a, b, h ,-, b ... totally 480 data, data primer obtained with second are b, a, h, a ,-... totally 480 data, totally 616 pairs of primers are added up respectively, and gained is the genotype data of this F2 colony.
With MapMaker3.0 software (ConstructinggeneticmapswithMAPMAKER/EXP3.0, SLincoln, MDaly, ELander-Cambridge, MA:WhiteheadInstitute, 1992) carry out genetic linkage maps drafting, obtain genetic linkage map. The position of 616 pairs of primers and the genetic distance with millet leaf color gene is can determine that from the genetic linkage map that this obtains.
(2) gene mapping
According to 480 individual leaf color phenotypes, similar to male parent type character is designated as a (blade is partially green), and similar to maternal type character is designated as b (the inclined yellow of blade), and character occupy and is designated as h between male parent and female parent. Obtain 480 individual phenotypic datas, 480 individual phenotypic datas are compared with 480 the individual genotype datas obtained before, similar Gao Ze represents this labelling and leaf color character close linkage, and by leaf color gene mapping on genetic linkage maps.
Embodiment 6: with the checking of the millet closely linked molecular marker of leaf color gene
1. on the basis of the genetic linkage maps prepared in embodiment 5, according to the genetic linkage distance with millet leaf color gene, the position being 0.5cM in the hereditary close linkage distance with millet leaf color gene determines molecular marker primer (SeqIDNo.2 and SeqIDNo.3), and find the male parent sequence location of correspondence, sequence between upstream and downstream primer is molecular marker, and its nucleotide sequence is such as shown in SeqIDNo.1.
SeqIDNo.2:5 '-GACATTCCAGTCGAGAAGCT-3 ';
SeqIDNo.3:5 ' GGTAGCTTCACTTCACAGAC-3 '.
2. additionally, in embodiment 5 in the electrophoresis of 480 individual pcr amplification products in F2 generation, amplification for molecular marker primer (SeqIDNo.2 and SeqIDNo.3) is: the amplified production of the plant that about 360 strain leaf colors are partially green all has the band of 731bp size, and the pcr amplification product of the plant of the about 120 inclined yellow of strain leaf color does not all have the band of 731bp (part amplification is as shown in Figure 1). And prove that through order-checking the sequence of the fragment of this 731bp is identical with SeqIDNo.1.
Visible, the molecular marker (SeqIDNo.1) of the present invention is and the millet closely linked molecular marker of leaf color gene.
Embodiment 7: molecular marker clone
The fragment expanding the 731bp of acquisition in embodiment 6 is cloned in pMD18-T carrier, it is thus achieved that recombinant vector. This recombinant vector is transformed in e. coli jm109, chooses monoclonal, cultivate and obtain reconstitution cell. Plasmid is extracted from reconstitution cell, described plasmid and recombinant vector, adopting M13 universal primer (sequence information is with reference to TaKaRa goods catalogue) that cloned sequence is checked order, result shows, contains the molecular marker (SeqIDNo.1) of the present invention in recombinant vector. The steps such as above-mentioned clone, conversion, cultivation, plasmid extraction are with reference to " the Molecular Cloning: A Laboratory guide third edition ", and yellow training hall etc. is translated, and Science Press's in JIUYUE, 2002 is published.
Above content is in conjunction with specific embodiment further description made for the present invention, it is impossible to assert that specific embodiment of the invention is confined to these explanations. For general technical staff of the technical field of the invention, without departing from the inventive concept of the premise, it is also possible to make some simple deduction or replace, protection scope of the present invention all should be considered as belonging to.
Claims (11)
1. one kind and the millet closely linked molecular marker of leaf color gene, it is characterised in that: described molecular marker is sequence shown in SeqIDNo.1.
2. that expand described in claim 1 and the millet closely linked molecular marker of leaf color gene a primer pair, it is characterised in that: the primer 1 of described primer pair is sequence shown in SeqIDNo.2, and primer 2 is sequence shown in SeqIDNo.3,
SeqIDNo.2:5 '-GACATTCCAGTCGAGAAGCT-3 ';
SeqIDNo.3:5 '-GGTAGCTTCACTTCACAGAC-3 '.
3. one kind and the millet closely linked molecular marker of leaf color gene, it is characterized in that: the primer pair that described molecular marker is described in claim 2 obtains through pcr amplification with the millet genomic DNA that blade is partially green for template, the millet of wherein said blade green partially is a paddy No. 1 and a confused flour beetle 3.
4. molecular marker according to claim 3, it is characterised in that: described molecular marker is sequence shown in SeqIDNo.1.
5. a carrier, it is characterised in that: containing the molecular marker according to any one of claim 1,3-4.
6. a reconstitution cell, it is characterised in that: containing the carrier described in claim 5.
7. the detection method of the molecular marker described in claim 1, including step: the nucleotide sequence design primer of molecular marker according to claim 1, expand for template with detected millet genomic DNA, and judge whether amplified production exists this molecular marker.
8. detection method according to claim 7, it is characterised in that: described primer is the primer pair described in claim 2.
9. claim 1, the molecular marker according to any one of 3-4 or the molecular marker primer pair described in claim 2 purposes in millet assistant breeding or millet leaf color gene mapping or detection.
10. the method for a millet leaf color gene mapping, it is characterised in that: described method includes the step using the molecular marker according to any one of claim 1,3-4 or the molecular marker primer pair described in claim 2.
11. a millet auxiliary breeding means, it is characterised in that: described method includes the molecular marker according to any one of test right requirement 1,3-4 or carries out the step detected by the molecular marker primer pair described in claim 2.
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| CN107385105B (en) * | 2017-09-22 | 2021-03-19 | 山西省农业科学院谷子研究所 | Molecular marker closely linked with millet glume color character, primer and application |
| CN108660235A (en) * | 2017-12-08 | 2018-10-16 | 深圳华大小米产业股份有限公司 | A kind of molecular labeling SIsv0686 with millet leaf color gene close linkage |
| CN108660234A (en) * | 2017-12-08 | 2018-10-16 | 深圳华大小米产业股份有限公司 | With the molecular labeling SIsv0362 of millet leaf color gene close linkage |
| CN113046458B (en) * | 2019-12-27 | 2023-05-23 | 深圳华大三生园科技有限公司 | Molecular marker closely linked with millet leaf included angle gene, detection method and application |
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| CN101974520A (en) * | 2010-11-22 | 2011-02-16 | 深圳华大基因科技有限公司 | Molecular maker SIsv1118 closely linked with high gene of millet strain |
| CN101974521A (en) * | 2010-11-22 | 2011-02-16 | 深圳华大基因科技有限公司 | Molecular marker SIsv0372 in close linkage with foxtail millet herbicide resistant gene |
| CN101982545A (en) * | 2010-11-22 | 2011-03-02 | 深圳华大基因科技有限公司 | Molecular marker SIsv0053 closely linked with millet plant height genes |
| CN101985621A (en) * | 2010-11-22 | 2011-03-16 | 深圳华大基因科技有限公司 | Molecular mark SIsvo188 closely linked with herbicide-resistant gene in foxtail millet |
| CN101985620A (en) * | 2010-11-22 | 2011-03-16 | 深圳华大基因科技有限公司 | Molecular marker SIsv1223 closely linked with millet herbicide-resistant gene |
Also Published As
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|---|---|
| CN102690811A (en) | 2012-09-26 |
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