CN107354232A - A kind of method for developing chromosome segment linkage molecule mark specific with wheat - Google Patents
A kind of method for developing chromosome segment linkage molecule mark specific with wheat Download PDFInfo
- Publication number
- CN107354232A CN107354232A CN201710842976.7A CN201710842976A CN107354232A CN 107354232 A CN107354232 A CN 107354232A CN 201710842976 A CN201710842976 A CN 201710842976A CN 107354232 A CN107354232 A CN 107354232A
- Authority
- CN
- China
- Prior art keywords
- wheat
- sequence
- mark
- molecular labeling
- exploitation
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- 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
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Analytical Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Immunology (AREA)
- Mycology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Botany (AREA)
- Biophysics (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
It is agricultural biological technical field the invention discloses a kind of method developed with specific chromosome of wheat section linkage molecule mark.It is the original series marked by the use of the EST in wheat deletion mapping, the ortholog of co-linear relationship determination, corresponding SNP as exploitation, the sequence that Wheat volatiles sequence that above sequence alignment is arrived and sequencing obtain is as the main sequence of exploitation mark, and Wheat volatiles sequence to acquisition and sequencing obtain sequence and carry out sequence signature analysis and marked so as to design corresponding SSR, STS, CAPS and dCAPS；The mark of above-mentioned exploitation is subjected to BSA analyses, genotyping and mapping analysis successively, obtains the molecular labeling with the specific chromosome segment close linkage of wheat.It is not high the present invention overcomes wheat molecular marker density, solve the problems, such as that wheat is relatively difficult in specific chromosome interval exploitation molecular labeling well.
Description
First, technical field
A kind of method for developing chromosome segment linkage molecule mark specific with wheat of present invention design, is specially that agricultural is raw
Thing technical field.
2nd, background technology
Wheat is that most important cereal crops, its Relationship with Yield are safe to world food in the world.Common wheat
(Triticum aestivum L.) is allohexaploid species, includes three different subgenomes (A, B, D).Genome surpasses
Cross 16 000Mb, containing repetitive sequence (Flavell etc., 1974, Genome size and proportion for having more than 80%
Of repeated nucleotide sequence DNA in plants, Biochem Genet, 12,257~269), molecule
Mark polymorphic rate low (Bryan etc., 1999, Low levels of DNA sequence variation among adapted
Genotypes of hexaploid wheat, Theor Appl Genet, 99,192~198).
With the development of experimental technique, the type of molecular labeling is gradually enriched, such as the RFLP (restriction at initial stage
Fragment length polymorphism, RFLP), AFLP (amplified fragment
Length polymorphism, AFLP) and according to sequence develop SSR (simple sequence
Repeats, simple repeated sequence), STS (sequence tagged sites, sequence tagged site), CAPS (cleaved
Amplified polymorphic sequence, digestion amplification polymorphism sequence), dCAPS (derived cleaved
Amplified polymorphic sequence, derivative digestion amplification polymorphism sequence) and SNP (single
Nucleotide-acid polymorphism, SNP) etc..The use of molecular labeling has greatly promoted new base
The excavation of cause, increasing disease-resistant, adversity gene identified by molecular labeling, such as Pm47 (Xiao M, 2013,
Identification of the gene Pm47 on chromosome 7BS conferring resistance to
Powdery mildew in the Chinese wheat landrace Hongyanglazi, Theor Appl Genet
126,1397~1403), Pm50 (Mohler etc., 2013, Pm50:a new powdery mildew resistance gene
In common wheat derived from cultivated emmer, J Appl Gene 54,259~63) and Pm51
(Zhan etc., 2014, Chromosomal Location and Comparative Genomics Analysis of
Powdery Mildew Resistance Gene Pm51in a Putative Wheat-Thinopyrum ponticum
Introgression Line, PLoS One 9, e113455) etc..
It is still relatively difficult in specific chromosome segment exploitation mark because wheat is also without complete genome sequence.
Generally require by the EST of physical positioning, co-linear relationship, genome sequence, SNP chip sequence information exploitation mark.Small
Mai Zhong, the heredity of Different Ploidy wheat and physical map information, the rice announced on the net, false bromegrass, a large amount of wheat est sequences letter
Breath and Wheat volatiles sequence information etc. are the vital signs sources for saturation target gene regions genetic map.These information
Mark to develop new is had laid a good foundation.The application patent proposes a kind of close in the specific chromosome segment exploitation of wheat
The method of linkage molecule mark.
In addition, booming high-flux sequence is also a kind of new strategy of genotyping in recent years.In order to reduce sequencing
Expense, researcher have been proposed some simplification based on two generation sequencing technologies represent library sequencing strategy (Hyten etc., 2010,
Highthroughput SNP discovery through deep resequencing of a reduced
representation library to anchor and orient scaffolds in the soybean whole
Genome sequence, BMC Genom, 11~38), wherein simplest method is to isolate and purify specific fragment magnitude range
Restriction fragment.Such as, using RAD, (restriction site-associated DNA, restriction enzyme site close
Join DNA) sequence measurement, successfully constructed using the method some dense genetic maps (Pfender etc., 2011, Mapping
with RAD(restriction-site associated DNA)markers to rapidly identify QTL for
Stem rust resistance in Lolium perenne, Theor Appl Genet 122,1467~1480；Wang
Deng 2012, Construction of a high-density genetic map for grape using next
Generation restriction-site associated DNA sequencing, BMC Plant Biol, 12~
148).And ddRAD (double-digest RAD, double digestion RAD) sequencings are a kind of sequencing strategy (Peterson derived from RAD
Deng 2012, Double digest RADseq:an inexpensive method for de novo SNP discovery
And genotyping in model and non-model species, PLoS One 7, e37135).DdRAD be sequenced with
RAD sequencings are similar, and compared with RAD single end sequencing, ddRAD double end sequencings can increase the specific and accurate of mark
Property.In recent years, in peanut (Davik etc., 2015, A ddRAD Based Linkage Map of the Cultivated
Strawberry, Fragaria xananassa, PLoS One 10, e0137746), strawberry (Zhou etc., 2014,
Construction of a SNP-based genetic linkage map in cultivated peanut based on
large scale marker development using next-generation double-digest
Restriction-site-associated DNA sequencing (ddRADseq), BMC Genomics 15,351) and oil
Dish (Wu etc., 2016, Evaluation of Linkage Disequilibrium Pattern and Association
Study on Seed Oil Content in Brassica napus Using ddRAD Sequencing, PLoS One,
11, e0146383) some dense genetic maps etc. are constructed using the sequence measurement in species.How in specific dyeing
Body section develops the molecular labeling of close linkage, and the application patent also provides a kind of utilize and simplifies sequencing strategy in specific dyeing
The method of body section exploitation compact linkage molecule mark.
The application patent proposes a set of combined method, i.e. exploitation develops linked marker with the specific chromosome segment of wheat
Method.
3rd, the content of the invention
Technical problem
The purpose of the present invention is directed to the deficiency of existing wheat molecular marker density, develops specific chromosome segment linkage molecule
Mark has difficulties, there is provided a kind of method of new exploitation chromosome segment linkage molecule mark specific with wheat.
Technical scheme
The technical scheme is that：
1. obtain the sequence information of exploitation molecular labeling：
1) according to the result of gene Primary Location, with reference to deliver SSR marker deletion mapping result (Sourdille etc.,
2004, Microsatellite-based deletion bin system for the establishment of
Genetic-physical map relationships in wheat (Triticum aestivum L.), Funct
Integr Genomics, 4,12~25), the chromosome arm where gene is primarily determined that, transfers and is positioned in the chromosome arm
Est sequence (Qi etc., 2004, A chromosome bin map of 16,000EST loci and distribution of
Genes among the three genomes of polyploid wheat, Genetics, 168,701~712；
http://wheat.pw.usda.gov), as original series；
2) false bromegrass genome (http is compared using the est sequence in above-mentioned chromosome arm://mips.helmholtz-
Muenchen.de/plant/Brachypodium/), rice genome (http://
Rice.plantbiology.msu.edu/), sorghum genome (http://mips.helmholtz-muenchen.de/
Plant/sorghum/ the synteny with the genome such as false bromegrass, rice, sorghum for) establishing the chromosome segment, find out and this
The wheat cdna sequence of a little genome orthologs, as original series；
3) according to the co-linear relationship of above-mentioned foundation, the co-linear relationship built with reference to 90kb SNP chips (Wang etc.,
2014, Characterization of polyploid wheat genomic diversity using a high-
Density 90,000single nucleotide polymorphism array, Plant Biotechnology
Journal, 12,787~796), estimation is located at the 90kb SNP markers of purpose chromosome segment, as original series；
4) utilize the trait segregation in segregating population to build the pond of two kinds of characters, respectively two kinds of ponds are carried out simplifying sequencing
(Hyten etc., 2010, Highthroughput SNP discovery through deep resequencing of a
reduced representation library to anchor and orient scaffolds in the soybean
Whole genome sequence, BMC Genom, 11~38), the SNP with trait associations of acquisition marks as design
Main sequence；
5) the SNP sequence alignment Wheat volatiles sequences obtained using the three kinds of original series and sequencing of above-mentioned acquisition
(http://www.wheatgenome.org/), the Wheat volatiles sequence that will be compared, the main sequence as design mark
(original series that generally, first three obtains can compare Wheat volatiles sequence；And what the 4th kind of sequencing obtained
The distinguished sequence of SNP sequences material sometimes, is compared less than Wheat volatiles sequence, and the SNP that acquisition is sequenced is also real anti-
Distinguishing base therein is answered, therefore, the 4th kind of sequence also serves as the main sequence of design mark).
2. the feature according to sequence is obtained develops different types of molecular labeling：
The Wheat volatiles sequence compared is subjected to signature analysis,
1) utilize in TANDEM REPEATS FINDER or SSR Finder software detection sequences whether include simple sequence
Repeat (SSR).If designing SSR marker in SSR both sides using primer-design software DNAMAN, if not provided, carrying out following the
2) step operates；
2) predictive genes website (http is utilized://www.softberry.com/) predict the gene that wherein whether there is
(especially Unigene), and be predicted to gene structure, using primer-design software DNAMAN gene 5 ', 3 ' and
Introne both sides design STS marks, if not having polymorphic in subsequent detection experiment, can be entered using corresponding restriction enzyme
Row digestion, CAPS marks are changed into, if without gene, carry out following 3) step operation；
3) SNP obtained using sequencing is converted into STS, CAPS or dCAPS mark.If there is the difference of restriction enzyme site in SNP
It is different, then the SNP marker is directly changed into CAPS marks, i.e., primer is designed in SNP site both sides, and utilized corresponding restricted
Restriction endonuclease carries out digestion；If the difference of restriction enzyme site is not present in SNP, (the http of software dCAPS Finder 2.0 are utilized://
Helix.wustl.edu/dcaps/dcaps.html) SNP is changed into dCAPS and marked with DNAMAN, i.e., is introduced in primer
Mispairing forms new restriction enzyme site with SNP site.
3. pair molecular labeling newly developed carries out BSA analyses：
Enter performing PCR in the mixing pit of two parents, two characters using above-mentioned molecular labeling newly developed to expand and utilize
Polyacrylamide gel (19:1、29:1、39:1) or Ago-Gel is (different according to clip size and the selection of difference size
Gel electrophoresis) that whether electrophoresis detection has is consistent polymorphic, and polymorphic carry out following 4th step if consistent and operate；If in two parents
This have it is polymorphic, do not have in the mixing pit of two characters it is polymorphic, then stop the mark continue operation (mark is likely to not exist
Purpose section)；If without polymorphic, SSR marker then stops operation, and STS marks then select corresponding restriction enzyme to carry out
Digestion, has seen whether consistent polymorphic, has, continues following 4th step operation, do not stop operation then.
4. showing consistent polymorphic molecular labeling in pair BSA analyses carries out genotyping, software Mapmaker is utilized
3.0b/EXP mapping analysis, determine molecular labeling newly developed whether with specific chromosome segment close linkage.
Further, the gene of molecular labeling to be developed is carried out BSA points according to step (1)~(2) step exploitation mark
Analysis, genotyping and mapping analysis, obtain the molecular labeling with the specific chromosome segment close linkage of wheat.
Further, during 3. step (2) the walks, if SNP has the difference of restriction enzyme site, directly the SNP is marked
Note changes into CAPS marks；If the difference of restriction enzyme site is not present in SNP, the Hes of software dCAPS Finder 2.0 are utilized
SNP is changed into dCAPS marks by DNAMAN.
Further, the acrylamide of the polyacrylamide gel described in step (3):Methene acrylamide is 19:1 or
29:1 or 39:1.
Further, molecular labeling clip size selects 19 in 100bp-150bp in step (3):1 polyacrylamide coagulates
Glue, molecular labeling clip size select 29 in 151bp-250bp:1 polyacrylamide gel, molecular labeling clip size exist
251bp-500bp selections 39:1 polyacrylamide gel.
Further, different gel electrophoresises is selected in step (3) according to molecular labeling clip size.
Beneficial effect
It is not high the present invention overcomes wheat molecular marker density, solve wheat well in specific chromosome interval
Develop the problem of molecular labeling is relatively difficult.By comparative genomics and existing reference gene group sequence exploitation mark, realize
The generality of the specific chromosome segment exploitation mark of wheat, pass through the mixed pond sequencing of packet and developed with existing reference gene group sequence
Mark, realize the particularity of the specific chromosome segment exploitation mark of Different Wheat Varieties.Compared with prior art, combine not
Same method, the generality of Wheat Cultivars exploitation mark was both considered, it is also considered that arrived Wheat Cultivars exploitation mark
Particularity.
This method is systemic, it is comprehensive it is strong and easy to operate, be easy to grasp.
4th, illustrate
Fig. 1 is the flow chart of exploitation chromosome segment compact linkage molecule mark specific with wheat.
Fig. 2 is exploitation and the genetic linkage mapses after pmHYM compact linkage molecules mark.
Fig. 3 is exploitation and the genetic linkage mapses after Pm48 compact linkage molecules mark.
5th, embodiment
Embodiment 1
With the exploitation of powdery mildew resistance gene in wheat pmHYM compact linkage molecules mark：
1. obtain the sequence information of exploitation molecular labeling：
1) according to pmHYM Primary Location result (Zhang Zhiliang master thesis), the gene positioned at mark Xgwm611 with
Between Xgwm577, due to (2008, the Microsatellite mapping of powdery mildew such as Nematollahi
Resistance allele Pm5d from common wheat line IGV1-455, Euphytica, 159,307~
313) mark Xgwm611 and Xgwm577 is positioned in wheat 7BL 0.86-1.00 chromosome arm, is located in the dyeing
The est sequence of body arm is as original series；
2) false bromegrass, rice, sorghum genome are compared using above-mentioned est sequence, establish the section and false bromegrass, rice with
And the synteny between sorghum, compare the co-linear relationship that 90kb SNP respective sections are built, it is determined that potential positioned at the section
SNP marker, as original series；
3) wheat 7BL genome sequences, the wheat 7BL genome sequences that will be compared are compared using above two original series
Row, the main sequence as marker development；
2. the feature according to sequence is obtained develops different types of molecular labeling：
The wheat 7BL genome sequences compared using TANDEM REPEATS FINDER or SSR Finder software detections
Whether include simple sequence repeats (SSR) in row.As a result show, SSR is included in the Wheat volatiles sequence compared, profit
SSR marker is designed in SSR both sides with primer-design software DNAMAN, develops 83 marks altogether；
3. pair molecular labeling newly developed carries out BSA analyses：
Enter performing PCR in two parents, anti-sense pond using above-mentioned 83 molecular labelings newly developed to expand and utilize polypropylene
Acrylamide gel (19:1、29:1、39:1) or Ago-Gel (selects different gel electricity according to clip size and difference size
Swimming) that whether electrophoresis detection has is consistent polymorphic, the results showed that and 12 marks therein show one between anti-sense parent and anti-sense pond
Cause polymorphic.
4. showing consistent 12 polymorphic molecular labelings in pair above-mentioned BSA analyses carries out genotyping, software is utilized
Mapmaker 3.0b/EXP mapping analysis, determine molecular labeling newly developed whether with specific chromosome segment close linkage.Figure
2 be exploitation and the genetic linkage mapses after pmHYM compact linkage molecules mark.Successfully obtained using the method for the application patent
5 are labeled as the molecular labeling of new close linkage with the molecular labelings of pmHYM more close linkages, overstriking.
Embodiment 2
With the exploitation of powdery mildew resistance gene in wheat Pm48 compact linkage molecules mark：
1. obtain the sequence information of exploitation molecular labeling：
1) according to Pm48 Primary Location result (Gao etc., 2012, Genetic analysis and molecular
Mapping of a new powdery mildew resistant gene Pm46in common wheat, Theor Appl
Genet, 125,967~973), the gene marks Xcfd81 and Xgwm205 may between mark Xcfd81 and Xgwm205
In wheat 5DS 0.63-1.00 chromosome arm, the est sequence of the chromosome arm is located in as original series；
2) false bromegrass, rice, sorghum genome are compared using above-mentioned est sequence, establish the section and false bromegrass, rice with
And the synteny between sorghum, compare the co-linear relationship that 90kb SNP respective sections are built, it is determined that potential positioned at the section
SNP marker, as original series；
3) by 50 pure and mild disease-resistant F2:3 familys are mixed into disease-resistant pond, 50 pure and mild susceptible F2:3 familys are mixed into disease-resistant
Pond, carry out simplifying sequencing, identification and the SNP sequences of trait associations, the main sequence as exploitation mark；
4) the sequence alignment wheat 5DS genome sequences obtained using above two original series and sequencing, comparison is arrived
Wheat 5DS genome sequences, as exploitation mark main sequence；
2. the feature according to sequence is obtained develops different types of molecular labeling：
1) the wheat 5DS genomes compared using TANDEM REPEATS FINDER or SSR Finder software detections
Whether include simple sequence repeats (SSR) in sequence.Include SSR in the Wheat volatiles sequence compared, utilize primer
Design software DNAMAN designs SSR marker in SSR both sides, develops 71 marks altogether；
2) it is being obtained using simplified sequencing to be converted into STS, CAPS or dCAPS mark with trait associations SNP, develop altogether
14 marks.
3. pair molecular labeling newly developed carries out BSA analyses：
Enter performing PCR in two parents, anti-sense pond using above-mentioned 85 molecular labelings newly developed to expand and utilize polypropylene
Acrylamide gel (19:1、29:1、39:1) or Ago-Gel (selects different gel electricity according to clip size and difference size
Swimming) that whether electrophoresis detection has is consistent polymorphic, the results showed that and 6 marks therein show consistent between anti-sense parent and anti-sense pond
It is polymorphic.
4. showing consistent 12 polymorphic molecular labelings in pair above-mentioned BSA analyses carries out genotyping, software is utilized
Mapmaker 3.0b/EXP mapping analysis, determine molecular labeling newly developed whether with specific chromosome segment close linkage.Figure
3 be exploitation and the genetic linkage mapses after Pm48 compact linkage molecules mark.Using the present invention method successfully obtain 3 with
The molecular labeling of Pm48 more close linkages, overstriking are labeled as the molecular labeling of new close linkage.
The present invention innovation have it is following some：
First, combined by being grouped mixed pond sequencing with comparative genomics, reference gene group, successfully solve different wheats
The problem of specific chromosomal region segment mark of variety development and particularity；
Second, this invention is more flexible, when wheat breed, which is applied to reference gene group exploitation mark, has general,
It can not have to carry out being grouped mixed pond sequencing, be developed when wheat breed is not suitable for traditional comparative genomics with reference gene group
Mark i.e. exploitation is marked when having particularity, then needs to carry out being grouped mixed pond sequencing；
3rd, the method for combining different type exploitations is systematic to propose in the specific chromosome segment of wheat
The method for developing compact linkage molecule mark.
Claims (6)
1. a kind of method for developing chromosome segment linkage molecule mark specific with wheat, its step are：
(1) sequence information of exploitation molecular labeling is obtained：
1. according to the preliminary positioning result of gene, the chromosome arm where gene is primarily determined that, transfers and is positioned at the chromosome arm
In est sequence, as original series；
It is somebody's turn to do 2. comparing false bromegrass genome, rice genome, sorghum genome using the est sequence in above-mentioned chromosome arm and establishing
Chromosome segment with false bromegrass, rice, sorghum genome synteny, find out the wheat-based with these genome orthologs
Because of sequence, as original series；
3. according to the co-linear relationship of above-mentioned foundation, with reference to the co-linear relationship of 90kb SNP chips structure, estimation is located at purpose
The 90kb SNP markers of chromosome segment, as original series；
4. building the DNA mixing pits of two kinds of characters using the trait segregation in segregating population, respectively two kinds of ponds are carried out simplifying survey
Sequence, by the SNP with trait associations of acquisition, the main sequence as design mark；
5. the SNP sequence alignment Wheat volatiles sequences obtained using the three kinds of original series and sequencing of above-mentioned acquisition, will be compared
The Wheat volatiles sequence arrived, the main sequence as design mark；
(2) different types of molecular labeling is developed according to the feature for obtaining sequence, the Wheat volatiles sequence compared is carried out
Signature analysis：
Whether 1. utilizing in TANDEM REPEATS FINDER or SSR Finder software detection sequences includes simple sequence repeats
SSR；If so, designing SSR marker in SSR both sides using primer-design software DNAMAN, 2. walked if not provided, carrying out following the
Operation；
2. the Unigene genes that wherein whether there is using the prediction of predictive genes website, and gene structure is predicted, utilize
Primer-design software DNAMAN is in 5 ', 3 ' and introne both sides of gene design STS marks, if do not had in subsequent detection experiment
Have it is polymorphic, can using corresponding restriction enzyme carry out digestion, change into CAPS mark, if without gene, carry out down
Face 3. step operation；
3. the SNP obtained using sequencing is converted into STS, CAPS or dCAPS mark；
(3) BSA analyses are carried out to molecular labeling newly developed：
Enter performing PCR in the mixing pit of two parents, two characters using above-mentioned molecular labeling newly developed to expand and utilize poly- third
Acrylamide gel or agarose gel electrophoresis detected whether it is consistent polymorphic, if consistent polymorphic progress step (4) step behaviour
Make；If having between two parents polymorphic, do not have polymorphic in the mixing pit of two characters, then stop the mark and continue to operate；
If without polymorphic, SSR marker stops operation, and STS marks select corresponding restriction enzyme to carry out digestion, have seen whether
It is consistent polymorphic, have and then continue below step (4) operation, do not stop operation then；
(4) consistent polymorphic molecular labeling is showed in analyzing BSA and carries out genotyping, utilizes software Mapmaker 3.0b/
EXP mapping analysis, determine molecular labeling newly developed whether with specific chromosome segment close linkage.
2. the method for exploitation chromosome segment linkage molecule mark specific with wheat according to claim 1, its feature exist
In by the gene of molecular labeling to be developed according to step (1)~(2) step exploitation mark, progress BSA analyses, genotyping
With mapping analysis, the molecular labeling with the specific chromosome segment close linkage of wheat is obtained.
3. the method for exploitation chromosome segment linkage molecule mark specific with wheat according to claim 1, its feature exist
In 3. the of, step (2) walks, if SNP has the difference of restriction enzyme site, the SNP marker is directly changed into CAPS marks
Note；If the difference of restriction enzyme site is not present in SNP, SNP is changed into using software dCAPS Finder 2.0 and DNAMAN
DCAPS is marked.
4. the method for exploitation chromosome segment linkage molecule mark specific with wheat according to claim 1, its feature exist
In the acrylamide of the polyacrylamide gel described in step (3):Methene acrylamide is 19:1 or 29:1 or 39:1.
5. the method for exploitation chromosome segment linkage molecule mark specific with wheat according to claim 4, its feature exist
In molecular labeling clip size selects 19 in 100bp-150bp in step (3):1 polyacrylamide gel, molecular labeling piece
Duan great little selects 29 in 151bp-250bp:1 polyacrylamide gel, molecular labeling clip size select in 251bp-500bp
39:1 polyacrylamide gel.
6. the method for exploitation chromosome segment linkage molecule mark specific with wheat according to claim 1, its feature exist
According to the gel electrophoresis that the selection of molecular labeling clip size is different in step (3).
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710842976.7A CN107354232A (en) | 2017-09-18 | 2017-09-18 | A kind of method for developing chromosome segment linkage molecule mark specific with wheat |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710842976.7A CN107354232A (en) | 2017-09-18 | 2017-09-18 | A kind of method for developing chromosome segment linkage molecule mark specific with wheat |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN107354232A true CN107354232A (en) | 2017-11-17 |
Family
ID=60291100
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710842976.7A Pending CN107354232A (en) | 2017-09-18 | 2017-09-18 | A kind of method for developing chromosome segment linkage molecule mark specific with wheat |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107354232A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108949925A (en) * | 2018-07-06 | 2018-12-07 | 中国水稻研究所 | A kind of molecular detecting method for quick and precisely identifying Weedy Rice and cultivated rice |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001027325A1 (en) * | 1999-10-08 | 2001-04-19 | Pioneer Hi-Bred International, Inc. | Marker assisted identification of a gene associated with a phenotypic trait |
| CN101760512A (en) * | 2008-11-19 | 2010-06-30 | 李祥 | RAPD marker research of wheat powdery mildew resistant gene Pm23 |
| CN102433327A (en) * | 2011-08-22 | 2012-05-02 | 江苏省农业科学院 | Molecular marker closely linked with wheat Tabasco powdery mildew resistance gene |
| CN105112416A (en) * | 2015-09-28 | 2015-12-02 | 江苏省农业科学院 | Molecular marker primer in close linkage with Hongyoumai powdery mildew resistance gene and application of molecular marker primer |
| CN106202995A (en) * | 2016-07-13 | 2016-12-07 | 北京麦美瑞生物科技有限公司 | The method of Semen Tritici aestivi BSR Seq gene mapping |
| CN106222285A (en) * | 2016-08-11 | 2016-12-14 | 江苏省农业科学院 | With powdery mildew resistance gene in wheat Pm48 be divided into from molecular marker |
-
2017
- 2017-09-18 CN CN201710842976.7A patent/CN107354232A/en active Pending
Patent Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2001027325A1 (en) * | 1999-10-08 | 2001-04-19 | Pioneer Hi-Bred International, Inc. | Marker assisted identification of a gene associated with a phenotypic trait |
| CN101760512A (en) * | 2008-11-19 | 2010-06-30 | 李祥 | RAPD marker research of wheat powdery mildew resistant gene Pm23 |
| CN102433327A (en) * | 2011-08-22 | 2012-05-02 | 江苏省农业科学院 | Molecular marker closely linked with wheat Tabasco powdery mildew resistance gene |
| CN105112416A (en) * | 2015-09-28 | 2015-12-02 | 江苏省农业科学院 | Molecular marker primer in close linkage with Hongyoumai powdery mildew resistance gene and application of molecular marker primer |
| CN106202995A (en) * | 2016-07-13 | 2016-12-07 | 北京麦美瑞生物科技有限公司 | The method of Semen Tritici aestivi BSR Seq gene mapping |
| CN106222285A (en) * | 2016-08-11 | 2016-12-14 | 江苏省农业科学院 | With powdery mildew resistance gene in wheat Pm48 be divided into from molecular marker |
Non-Patent Citations (5)
| Title |
|---|
| BISHENG FU ET AL: "Identification and mapping of a new powdery mildew resistance allele in the Chinese wheat landrace Hongyoumai", 《第八届全国小麦基因组学及分子育种大会摘要集》 * |
| 付必胜 等: "与小麦抗白粉病基因Pm48紧密连锁分子标记的开发", 《作物学报》 * |
| 刘颖 等: "小麦纹枯病抗源的遗传多样性及抗性基因位点SSR标记分析", 《作物学报》 * |
| 张志良: "小麦地方品种红蚰麦抗白粉病基因的发掘和定位", 《中国优秀硕士学位论文全文数据库 农业科技辑》 * |
| 曹墨菊 等: "《植物生物技术概论》", 31 October 2014, 中国农业大学出版社 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108949925A (en) * | 2018-07-06 | 2018-12-07 | 中国水稻研究所 | A kind of molecular detecting method for quick and precisely identifying Weedy Rice and cultivated rice |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Nadeem et al. | DNA molecular markers in plant breeding: current status and recent advancements in genomic selection and genome editing | |
| Adhikari et al. | Application of molecular markers in plant genome analysis: a review | |
| Parida et al. | Informative genomic microsatellite markers for efficient genotyping applications in sugarcane | |
| Saxena et al. | An integrated genomic approach for rapid delineation of candidate genes regulating agro-morphological traits in chickpea | |
| Fernández-Silva et al. | Bin mapping of genomic and EST-derived SSRs in melon (Cucumis melo L.) | |
| Xu et al. | A high-density genetic map of cucumber derived from Specific Length Amplified Fragment sequencing (SLAF-seq) | |
| Chapman et al. | A genomic scan for selection reveals candidates for genes involved in the evolution of cultivated sunflower (Helianthus annuus) | |
| Stephens et al. | Genetic marker discovery, intraspecific linkage map construction and quantitative trait locus analysis of Ascochyta blight resistance in chickpea (Cicer arietinum L.) | |
| Cogan et al. | Gene-associated single nucleotide polymorphism discovery in perennial ryegrass (Lolium perenne L.) | |
| Xu | Global view of QTL: rice as a model. | |
| Wanchana et al. | A rapid construction of a physical contig across a 4.5 cM region for rice grain aroma facilitates marker enrichment for positional cloning | |
| Kujur et al. | An efficient and cost-effective approach for genic microsatellite marker-based large-scale trait association mapping: identification of candidate genes for seed weight in chickpea | |
| Jia et al. | Marker development using SLAF-seq and whole-genome shotgun strategy to fine-map the semi-dwarf gene ari-e in barley | |
| Varshney | Gene-based marker systems in plants: high throughput approaches for marker discovery and genotyping | |
| Han et al. | QTL mapping pod dehiscence resistance in soybean (Glycine max L. Merr.) using specific-locus amplified fragment sequencing | |
| CN110923352B (en) | KASP marker of wheat powdery mildew resistance gene PmDTM and application thereof | |
| Yang et al. | Methods for developing molecular markers | |
| CN107090495A (en) | The molecular labeling related to the long character of millet neck and its detection primer and application | |
| CN107090494A (en) | The molecular labeling related to millet code grain number character and its detection primer and application | |
| Maloof | Genomic approaches to analyzing natural variation in Arabidopsis thaliana | |
| Sandhu et al. | Development and validation of a novel core set of KASP markers for the traits improving grain yield and adaptability of rice under direct-seeded cultivation conditions | |
| Bohra et al. | Translational genomics and molecular breeding for enhancing precision and efficiency in crop improvement programs: some examples in legumes | |
| Drost et al. | A microarray‐based genotyping and genetic mapping approach for highly heterozygous outcrossing species enables localization of a large fraction of the unassembled Populus trichocarpa genome sequence | |
| CN107090450A (en) | The molecular labeling related to millet spike length character and its detection primer and application | |
| US6368806B1 (en) | Marker assisted identification of a gene associated with a phenotypic trait |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |