CN109517925A - Flax SSR molecular marker and its application - Google Patents
Flax SSR molecular marker and its application Download PDFInfo
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- CN109517925A CN109517925A CN201910067537.2A CN201910067537A CN109517925A CN 109517925 A CN109517925 A CN 109517925A CN 201910067537 A CN201910067537 A CN 201910067537A CN 109517925 A CN109517925 A CN 109517925A
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- 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
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- 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
Abstract
The present invention relates to plant variety identification and breeding technical fields, and in particular to the construction method of flax SSR finger-print.The present invention provides through MISA softwares to obtain 28751 sites SSR to flax genome search, devises 71184 pairs of primers altogether using Primer 5.0, utilizes 26 parts of Flax Germplasms, screen the SSR marker of 11 pairs of rich polymorphisms: RM4-5, RM4-7, RM6-2, RM6-6, RM6-7, RM7-1, RM7-3, RM8-1, RM9-4, RM13-5 and RM15-1.SSR primer of the present invention based on the exploitation of flax genome have many advantages, such as amplification stablize, electrophoretic band clearly, rich polymorphism, the present invention can be effectively used for the research fields such as Flax Germplasm analysis of genetic diversity, the building of high density map, variety and authenticity identification, molecular mark.
Description
Technical field
The present invention relates to plant variety identification and breeding technical field, in particular to flax SSR molecular marker and its applications.
Background technique
Flax (Linum usitatissimum L.) is a kind of important industrial crops, is widely cultivated in northwest China
The Northeast and.The fiber that flax stem is produced is the important source material of textile industry, flex seed oil content 30%-45%, flax
Oil is widely used in the industries such as oil expression and health care rich in unsaturated fatty acids such as linolenic acid, linoleic acid.Due to the fiber of high-quality
With the shortage of linseed, the further development of Fiber Flax Industry is limited.Forefathers are studies have shown that molecular breeding is crop improvement quality
One of the important means of, and molecular markers development is one of molecular breeding important step.Although the molecule of some based on PCR segments
Label such as RAPD, AFLP, ISSR are widely utilized from flax is studied, but the generally existing time-consuming, polymorphism of these labels
The features such as low (Yurenkova et al., 2005；Fu,2006；Everaert et al.,2001).
Microsatellite (Microsatellite) refer to wide dispersion in genome with a few nucleotide, (majority is
2-4) be unit multiple tandem sequence repeats DNA sequence dna, people are referred to as simple repeated sequence (Simple
Sequencerepeats, SSR), it is the relatively high a kind of DNA sequence dna of aberration rate in genome.In recent years, SSR has become
Most popular genetic marker.As the higher codominant marker of detection efficiency, SSR marker has concentrated the excellent of other molecular labelings
Point, such as typical codominance and multiple alleles, have high polymorphism, can accurately distinguish affiliation very similar
Body；The dosage of DNA is few, and the reproducibility of amplification is high, can effectively realize simple, quick PCR operation, while genotype
It can also can be divided automatically by the fluorescent marker of primer by agarose gel electrophoresis and Polyacrylamide Gel Electrophoresis
Analysis.In recent years, having had relevant report about the identification in the site SSR and the exploitation of SSR molecular marker in flax gene
(Cloutier et al.,2009；Cloutier et al.,2011；Soto-Cerda et al.,2011；Cloutier et
al.,2012；Wu et al.,2017).Cloutier etc. (2009) is from 146611 ESTs from 10 libraries
851 sites SSR are identified, devise 662 pairs of SSR primers altogether.It is come out although some sites SSR are identified, while corresponding
SSR marker is developed, but still remain that SSR marker number is on the low side, mark density is low and the physical location information of label not
The disadvantages of knowing, it is difficult to meet the building of high density linkage map, the finely positioning of gene, flax molecular breeding etc..Flax
The sequencing of full-length genome is to develop a large amount of SSR markers to provide the foundation (Wang et al.2012；You et al.,2018).And
And the sequencing of the full genome of high quality in the recent period is not only that researcher provides accurate sequence information, and the physical bit refined
Confidence breath is that the Molecular and Genetic Study of flax lays the foundation.The searching in the site flax full-length genome SSR, SSR molecular marker are opened
Hair has no relevant report.
Summary of the invention
In view of this, the present invention provides flax SSR molecular marker and its applications.The present invention utilizes flax gene in NCBI
Group sequence information develops flax microsatellite molecular marker using bioinformatics method, and utilizes 96 pairs of micro-satellite molecule marks
Note genetic affinity analysis has been carried out to 26 parts of linen knitting yarns, obtain amplification stablize, band it is clear, 11 pairs of rich polymorphism
SSR marker.These labels can be effectively used for genetic analysis, the high density map construction of Flax Germplasm.
In order to achieve the above-mentioned object of the invention, the present invention the following technical schemes are provided:
The present invention provides flax SSR molecular marker, flax SSR molecular marker has such as No.1~11 SEQ ID
Any one of or appoint several shown in sequence.
It is high in Flax Germplasm analysis of genetic diversity, flax that the present invention also provides flax SSR molecular markers
Application in the building of density map, Flax Varietieies identification and/or molecular mark.
On the basis of the studies above, the present invention provides the primer sets for expanding flax SSR molecular marker, including
(1) several groups of primers are appointed in~any group of (11):
(1), expand the upstream primer of the sequence as shown in SEQ ID No.1: its sequence is as shown in SEQ ID No.12；
Expand the downstream primer of the sequence as shown in SEQ ID No.1: its sequence is as shown in SEQ ID No.13；
(2), expand the upstream primer of the sequence as shown in SEQ ID No.2: its sequence is as shown in SEQ ID No.14；
Expand the downstream primer of the sequence as shown in SEQ ID No.2: its sequence is as shown in SEQ ID No.15；
(3), expand the upstream primer of the sequence as shown in SEQ ID No.3: its sequence is as shown in SEQ ID No.16；
Expand the downstream primer of the sequence as shown in SEQ ID No.3: its sequence is as shown in SEQ ID No.17；
(4), expand the upstream primer of the sequence as shown in SEQ ID No.4: its sequence is as shown in SEQ ID No.18；
Expand the downstream primer of the sequence as shown in SEQ ID No.4: its sequence is as shown in SEQ ID No.19；
(5), expand the upstream primer of the sequence as shown in SEQ ID No.5: its sequence is as shown in SEQ ID No.20；
Expand the downstream primer of the sequence as shown in SEQ ID No.5: its sequence is as shown in SEQ ID No.21；
(6), expand the upstream primer of the sequence as shown in SEQ ID No.6: its sequence is as shown in SEQ ID No.22；
Expand the downstream primer of the sequence as shown in SEQ ID No.6: its sequence is as shown in SEQ ID No.23；
(7), expand the upstream primer of the sequence as shown in SEQ ID No.7: its sequence is as shown in SEQ ID No.24；
Expand the downstream primer of the sequence as shown in SEQ ID No.7: its sequence is as shown in SEQ ID No.25；
(8), expand the upstream primer of the sequence as shown in SEQ ID No.8: its sequence is as shown in SEQ ID No.26；
Expand the downstream primer of the sequence as shown in SEQ ID No.8: its sequence is as shown in SEQ ID No.27；
(9), expand the upstream primer of the sequence as shown in SEQ ID No.9: its sequence is as shown in SEQ ID No.28；
Expand the downstream primer of the sequence as shown in SEQ ID No.9: its sequence is as shown in SEQ ID No.29；
(10), expand the upstream primer of the sequence as shown in SEQ ID No.10: its sequence is as shown in SEQ ID No.30；
Expand the downstream primer of the sequence as shown in SEQ ID No.10: its sequence is as shown in SEQ ID No.31；
(11), expand the upstream primer of the sequence as shown in SEQ ID No.11: its sequence is as shown in SEQ ID No.32；
Expand the downstream primer of the sequence as shown in SEQ ID No.11: its sequence is as shown in SEQ ID No.33.
The present invention also provides the primer sets to expand the application in flax SSR molecular marker.
The present invention also provides the primer sets in Flax Germplasm analysis of genetic diversity, flax high density map
Application in building, Flax Varietieies identification and/or molecular mark.
The present invention also provides kits, including the primer sets.
The present invention also provides the kits in Flax Germplasm analysis of genetic diversity, flax high density map
Application in building, Flax Varietieies identification and/or molecular mark.
The present invention also provides the methods of Flax Germplasm analysis of genetic diversity, using the primer sets to flax
Sample to be tested is expanded, electrophoresis detection, obtains genotype data；
The genotype data reads standard are as follows: same group of primer pair difference flax sample to be tested amplification, electrophoresis detection knot
In fruit, maximum band is denoted as A, is secondly B, and so on, band is unintelligible or missing is denoted as 0；Utilize PowerMarker
3.25 obtain the number of the PIC value of every group of primer and allele, obtain Flax Germplasm genetic diversity according to the PIC value
Property result.
The present invention also provides the methods of Flax Varietieies identification, are expanded using the primer sets flax sample to be tested
Increase, electrophoresis detection, obtains genotype data；
The genotype data reads standard are as follows: same group of primer pair difference flax sample to be tested amplification, electrophoresis detection knot
In fruit, maximum band is denoted as A, is secondly B, and so on, band is unintelligible or missing is denoted as 0；Utilize PowerMarker
3.25 obtain the number of the PIC value of every group of primer and allele, obtain Flax Varietieies qualification result.
The present invention also provides the preparation method of flax germplasm dendrogram, using the primer sets to flax sample to be tested into
Row amplification, electrophoresis detection obtain genotype data；
The genotype data reads standard are as follows: same group of primer pair difference flax sample to be tested amplification, electrophoresis detection knot
In fruit, maximum band is denoted as A, is secondly B, and so on, band is unintelligible or missing is denoted as 0；Obtain every group of primer
Genetic distance carries out clustering to Flax Germplasm based on the genetic distance.
The present invention is using flax Genomic sequence information in NCBI, using bioinformatics method, develops that flax is micro- to be defended
Star molecular labeling, and genetic affinity analysis has been carried out to 26 parts of linen knitting yarns using 96 pairs of microsatellite molecular markers, expanded
Increasing is stablized, band is clear, 11 pairs of SSR markers of rich polymorphism.These labels can be effectively used for Flax Germplasm
Genetic analysis, high density map construction and the assignment of genes gene mapping.
Detailed description of the invention
In order to more clearly explain the embodiment of the invention or the technical proposal in the existing technology, to embodiment or will show below
There is attached drawing needed in technical description to be briefly described.
Fig. 1 SSR primer RM4-5 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Fig. 2 SSR primer RM4-7 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Fig. 3 SSR primer RM6-2 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Fig. 4 SSR primer RM6-6 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Fig. 5 SSR primer RM6-7 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Fig. 6 SSR primer RM7-1 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Fig. 7 SSR primer RM7-3 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Fig. 8 SSR primer RM8-1 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Fig. 9 SSR primer RM9-4 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Figure 10 SSR primer RM13-5 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Figure 11 SSR primer RM15-1 expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Figure 12 SSR primer (negative control) expands the silver staining PAGE figure of 26 parts of flax germplasm DNA samples；
Figure 13 is based on above-mentioned SSR primer amplification result 26 portions of flax germplasm dendrogram obtained.
Specific embodiment
The invention discloses flax SSR molecular marker and its application, those skilled in the art can use for reference present disclosure, fit
When improvement realization of process parameters.In particular, it should be pointed out that all similar substitutions and modifications are for a person skilled in the art
It is it will be apparent that they are considered as being included in the present invention.Method of the invention and application passed through preferred embodiment into
Gone description, related personnel obviously can not depart from the content of present invention, in spirit and scope to method described herein and application
It is modified or appropriate changes and combinations, carrys out implementation and application the technology of the present invention.
The present invention is using flax Genomic sequence information in NCBI, using bioinformatics method, develops that flax is micro- to be defended
Star molecular labeling, and genetic affinity analysis has been carried out to 26 parts of linen knitting yarns using 96 pairs of microsatellite molecular markers, expanded
Increasing is stablized, band is clear, 11 pairs of SSR markers of rich polymorphism.These labels can be effectively used for Flax Germplasm
Genetic analysis, high density map construction and the assignment of genes gene mapping.
To realize that above-mentioned target, the invention patent scheme are as follows:
(1) feature of flax SSR molecular marker includes: the exploitation of SSR molecular marker；The title of SSR molecular marker；
The primer sequence of SSR molecular marker；SSR molecular marker on reference genome physical location (https: //
www.ncbi.nlm.nih.gov/assembly/GCA_000224295.2)；Using Flax Varietieies CDC Bethune DNA as mould
The PCR product clip size of plate amplification.
SSR marker exploitation: NCBI first download flax whole genome sequence information (https: //
Www.ncbi.nlm.nih.gov/assembly/GCA_000224295.2), MISA software is further utilized
(MIcroSAtellite identification tool) (http://pgrc.ipk-gatersleben.de/misa/) inspection
Survey the microsatellite in flax genome sequence and compound microsatellite locus.The detection parameters used is 2-6,3-5,4-5,5-5,6-
5 (number of repetition of 2 base repetitive unit motif >=6 times；The number of repetition of 3 base repetitive units >=5；And so on.)；
When the distance between two microsatellites are less than 100bp, then a compound microsatellite is formed, is examined in entire flax genome
Measure 28571 sites SSR.According to the SSR testing result of MISA, Primer3 (http://pgrc.ipk- is utilized
Gatersleben.de/misa/primer3.html) software default parameter (length 100bp-300bp；55 DEG C -60 DEG C of Tm value,
Primer length 20-25bp) design of primers is carried out, the highest 3 pairs of primers of marking are selected in design result is used for subsequent experiment,
71184 pairs of SSR primers are obtained.
The title of SSR molecular marker are as follows: RM4-5 (nucleotide sequence is as shown in SEQ ID No.1), RM4-7 (nucleotides sequence
Column are as shown in SEQ ID No.2), RM6-2 (nucleotide sequence is as shown in SEQ ID No.2), RM6-6 (nucleotide sequence such as SEQ
Shown in ID No.4), RM6-7 (nucleotide sequence is as shown in SEQ ID No.5), RM7-1 (nucleotide sequence such as SEQ ID No.6
It is shown), RM7-3 (nucleotide sequence is as shown in SEQ ID No.7), RM8-1 (nucleotide sequence is as shown in SEQ ID No.8),
RM9-4 (nucleotide sequence is as shown in SEQ ID No.9), RM13-5 (nucleotide sequence is as shown in SEQ ID No.10) and
RM15-1 (nucleotide sequence is as shown in SEQ ID No.11).
SEQ ID No.1:
TTGGTCGTGTGTACGGAGTTAATATGGAAAATACACGCAGAGATTTGGATACGATTTTAGGTTTCTAC
CTTCCTATCTCATACTCACGTGTCGTTACAACAATGTCATTTGGTATAGTGCTAActaaaaagttcaattttttta
ttataaatagctCACAAATCTGGACGATCTTAAAGACAGGACTATCGACTCCATAGCAATTTCATATCGAAAGACG
GAGGATGAGATAGGACCAAAGTCATACTAACCTACAAAATCCCAATTTCCATTTTTGCTCTTTTACTGAGATATGT
ACacttgaatttcaattttgacatagTTTAAACAgtaatttttcaaacttttaaagggaaaacataaattttacaa
acttTTAAAGGAAAAAGTAATGGGAGGTAAAGTTGAAAGGAACACAGGAATTGATGTATGTGAAAATTTGCCATGA
TAACAGGGAACATTGGTGCATGAAGGGTCTCGATCTTTGCATACAGaagcaaataataataataataataataata
ataatctctTCTCATGTGACATTGGGAATAATAATCGTAAGATTTAATACGAGTATGTCTCTTTATTTACAGATCT
ACACTATCATTACCCCAATGGGGTTGTGCTATGTTTTGACCTATGAACCATCTTTCATCTCGTAGCGGCGATAGAA
GTGAGAAAAAcgtaattaaaaatgttagaTTAATAACGAGAGTGTGTGAATTACAAAATGTGAAAGGACAGTCTCA
TTTAAATTATCATGTAAAGATTATTAGGAATCGtcgaaaaatcaaagtttaaaCTATTGAATGTctaaaatgacaa
aatgatGCACATTTACGCGATTTTGGAAGGCTCGATGATGCTACAACATCTCGCAAGCTAATCTCCTAGTGATTCT
CTTATGCATCTATATACCACAACGAAACttatcatgttgtctttcattaaccactaatttttttatgtgactACTT
ACTTACTAGGTTGGGGATGG
SEQ ID No.2:
AAGGTCAGTCCCACGTTCATCTCCTTGGCTGCCTCCGTTATCGGCATGTAGAAATACCTCGACAGCAT
TTCCCTCGTGATCACCTGCTGCCCGTTTGGCGAAACCGCTTTGTTATGCTCATTCGTTGTTGTTTTCTGTTCATGA
TCAAGGATTGGGAGGAGAGGTGGCCCCAGTTCGTTGCAAACCGGATCGTGATCGATGTGGGAAGaacatgaagatg
atgatattgatgaagatgacgaatgatgaagaatattaGTACTActgatcatcatattcatgatGTCAGTACCGTA
GCGATCACCATCGGCAGCATTAACTATATCGGGATCTTGGTAGTATTGATCAAGCTGGGTAAAAGGTGCTTCGAGT
TCGAACTGATGATTGAATAATGGGAATGGAggaagggatgatgatgagtagtAGTCGTCATCCCCACCACCATGGC
TGCTGTACAGACACATATATAGAAAGAgggaatattattaattatgatggcgatgatgatgatgatgtgtaagtaa
ttaattaacgtaCGATATTACCCAAAAGTGAAGTCGTCAAAGAGTCCTCCGAATGGCGCTTGTTGCTGGGAATCGT
CATGGAGAAGTGGCGGTGATGAACGGATTTGGAAGTGATCTTGTTCTTGCTTCACCTCAACAAACTCcattgtttg
atgatgatgatgatgacgatgagaACACAGAGCCAAATGAAGGGAGGGGGAGTGGTTTATATGTAGAGTGcgggtg
atgaatgatgattgtGTGCAGTAAAAAATCTAGTGGATTAAAAAAGCTGAGAAAACTATTAGGGGAAGTCGATGGG
TTGCTGTTTTGGAGGAGATGCATAATTACAGTGAGAGACGGAAAGGGGAGTTActttttttacatattggATTAGT
GATagtgattaataattaatatcgGAGTAGTAATTGCAGTAAttcattgtaattttaatGGGGGGAATTCATTATT
GATGTCAATATAATTAACAC
SEQ ID No.3:
CACTTGGTCGATAGCGTCCTCCGATACTCTGCCTCCGCCAATCCGGACGGAGCCGGAAGAAAGACCAG
CTGGAGCCGAAACCAAATCATCAGGTAGTTCCAGCTGAGTCGATTGTAGCTTGGAGTCCTTGAGCTTTAAACGATT
CATAGTATCGTCCAAATCGGCCAACTCCCATGAATCCGGGGCTCCGAGATCATCTGACACAGAGCCCTCCATGGAT
TCTTTGACGCAGAGAGCTAACGGAATAAGGAGGTGAACGCTAAGTCACCTGAAAAGGAACCTTAAAGTGCAAATCA
ATCGTCGCTAGGGTTGTCGACGTTCCAGCAATATTGTGTTGAGCATCGTGTTATATGACCGTAAATCGACCGATCC
AGGATTGCGATTAGGCCGACGACTCCGAGGCGTGAATCACATGCAGAATGCGACTCGtagggttagggtttcgaAT
TCCACCGGATTATTTGGCTAAAATTGGAGAGGCCTACAATCTTTCTTacatagagagagagagagagagagagagg
gagaaagagagagagagagaatgagAGAGGATAGAGTTTGATAAGTCGTTTGTCGAAATTCGAAAGGGGGTGTGTG
TAGCGTCTCGTATCGTATGCGCTTAAGCACAGTAGAAAAAACGAAAGGTcggaaatttaaaatataagagatttaa
ataaattttattatttttaaaataatttataagaaattttaaatatgattaTTAGTGTAGTAATCTATTGAtaagt
tgataatttgattagtAATAATTGTATAATAAGCCAATTTAGTTTAGCAGAatgcgaattttattagaaataaatt
aaatagtgttttatttgtttaaatgtagttgttgaaaaaaatggcgaagacaaaGGGTGGAGCTAAATCGTACAAT
TAagatccggtagaggaagaaacgaatggaattcagaaaatgagagaaaaaggaaagcaagcTGGCATTTTAAATC
GAATGAAAGAGCGAGTGATC
SEQ ID No.4:
aaacagtgacgctttcttccaaagtcgaacgctttgagttgttccggtcTCCTCATCATGTtttggtt
tatctccaaacgtcatatgacacctgaaatataccaaaaacataccaaacaccataataccgccccgaggggcaag
agaaccaaacatgtactcacaacccaaatttgaatacatttcgacacacaaaaatatagtttataacgagaaaagg
tatgtcgaaaatacatcacatcactGCTAACTACAATAACACTTAATTAGTAATGACCACAACTTgatttttcacc
tttttttcaaaaggCAAATACTTCAACTTTgataaaaatgaatcaaaattcagCCTCTCGACTAATAACATCAGAA
ATCTCAAAAGCCATTAAATTTCAATGAGCAAGTCTATTTGCTTCTCGACAGACAAAACTCCAATCTCAAATATCAC
AACGAGCAATTCCACTCGACAGACGCTGCCAGCTTCAGTCCTGCAaatatccttcttcttcttcttcttcttcttc
agcaaatCAATAAGCTTTTTACAGTCCAATTTTTGAACTTAAAAATGGGGGAGAGTgtgttcaaaattaaattaaa
gacAAAAAGCCTCAACATAATTCTTGTTGTTTGAGTGAGAGTGCGTGTGGTTTTCGGAGTGTTTGCATGATATGAT
ATAATATGATATGACcccatccatcttcttcctccctcaatatcatataaaatcaaagtttctttctttctttccc
cttttcccttttcatctctctcctctctctctatctctctgtACTCTGtgtctctttctttttcagctgcttcttc
ttcttcttcttcttcttcttcctggtgATGGCTTTTTCATCTGTACCTCTGTCGGCCATCGATTTCCTTTCCATTG
AATCTTTGATATCCACATGAAAGAAGATTCCTTCGCTGCctactttttttctcttgattttccttcttctcccccC
TAAGGTCATGTGATTCAGGC
SEQ ID No.5:
TTCCAGCTGTCCCACGTGTCGTGCATCCATGATCCCAGTAGGTCCTTTGTCGACGTGTGCCAGACGGG
CTGACGTGGAAGCTTACAGTGTACGAGGCAATATGGTTGGAGATGAATTTGGAGTTGCTCGATAATGTAAAATTGA
TTCGAcgatataattataattttatgaacaATTGGTGCATTATTAAGATTCCTCTGTGTCCTCTATTTCCCCCCCA
TATTTTAACCCAGAGATGGATGGGGAGAATTATGTGCAAGAAATGAAGggtgaagaaaagaatatgatGTGTTGAG
CATGTGAATATACAAAGTCATGTAAATAGAAAGACGATTCCTGTTTAGGAAGCAAAGAAAGGTGCTCGACGGGGaa
tcccttttcttcttcttgcgTTAGAAGATGGAATTGCTTTGTACATATCTTCTCCTTGTTGCAATTCATGTTCATT
TGGTGATGACGATGACGACATCATCGCTCTAGACCTCCCTAtaactcttctttcttcttcttcttcttcttcctcg
tttgcttcttcttctccccttccttcaTTCTTGCCTTCCTTGCTAAACCTTGCATTCCCATTCTTCTTTATTCCAA
CAAGCTACAAGGACATAACCATGTAATTAATTCCCCACTAATTAACTTGTTATCACATTTGGTCAAAACCGTATTA
TATTACCACTCGGCCAATTCCTTGGAGTTCTGAATTCGGACACTTTAGTAACTTATACCGTAAAGTGTCATTaggt
aaattatttttcaatccaaCACATGCATTGATAATCTTGATTTCCATAGTCAACTTTTTGAAAGATGGAAAACCAA
CCataggtgatgatgatgaagcttgACGTATTAAAAACATCATTTGGGTTaggtattttatatatgtagtATTTTG
AAACATTATTTAAAGATATGGTATTTATGAGATAAAATTTGTGATGTGTAAATTATCTTGTTTAACTATTCAACGTG
GTAGTTTATCGGGGTATTT
SEQ ID No.6:
TACAAATTCGGTGTGGGTCGATAATGGACAGCTGACTCATATCTAGTCGAATTAGTCCAAGCAGGTCG
AGTTTGAATTGTCAtttgagaggaagagaatcaAATGGAGGATTCATCAATATACACCACTTCTTTACATTgaata
acaaactcaaactcttggggcacaaaaaaatttgggaaGAGATCGAGTTCATTCATCTCTTTGAACTCAATATAAA
GACTGTTCGAATCTTGAGACCAACAATCATCTATTATCTAACAGGCATAGGGAGCGAAAACCCTAACAGCTGAGCG
CGGAGGTGAGACTAACGAAGTACTCCTCGCTGCACGGGATGGTGAGTCCGCCCATGGGGTGATCGAACCCGAACTC
TTCCTCGGCTCGATCGAGTAAGACTTGGAACAAAGGGTGGTTCAAGTAAGTGACTGGGATGAtgaacctcttcttc
ttaagGTAGCCTTCACCAACATAGACTGCAATATGGCCCTTTGGCacattggaagaagaagaagaagaagatagtt
TGGCTGACAGTGTTCTTTGTAGCTTTTGCCTTGCATTTTCAGCAATCCCCATTGTTAAGAAACAGAGgttgagaaa
cagaggaaaagaaggagtTTTTATTGgtgggttttgagtttttactAAGTTTGTTTCATGCATGAAGGcttatata
tagagagaggtGGGGTTTTGCAAGTGAAATTCTTGAAAGAATAATTgtaagtagaagaagaagaagaaacatacCA
GGTGAAGAGGCTGCTGAAGATTCACATGGGTGTGCTTGTGTCCATGGCTACATCCTCTGGCCTACTGTCTTTTTCAG
TTGAGTGCTATGAGCTACATATTCTGTGTTCACATAAGAGTTCAAAGTTTGCTTATCTAAGCAAGGCAAGGGGTTTA
ACTACGTTCAATTTCGTTATTCAGCCACACCCTAACCGACAGTGCCCTCAACTGAGTGGCTTTAGGCACAACGTGTT
GCATAGAAGTAGCTTGA
SEQ ID No.7:
AGGCGGGTTGATCAGGTTAACATTTATAGGGGTTCAATCATGTGACATTTGTTTGTATTCGGGTTGGG
GTTGGGTTGGGGGGGGAGGTTATTAGATTAATGGCTTCTAGTCGATTCAAATCATTAACCGAGTCAGATAAGACTC
GAACTGAGTGGATGCAAGATCAGCTTCTTAATTACAATTCTTATTGTATAATCCACATTAACTGATTAACGGTGGT
TGGTTGATTGCAGATGTACCGTACGGTAAAGACCACTGACAGAGCTGCTGCCACGTCAGGTTAACGACTTAACCCC
CAAAATGAACTTACTTATATGATCAATATCAATGGTTGCTTCATGATATTTACGGAAATACTTTTGCAACATATGT
TTACAGGGCAATCAGATTTAGTTGACAACGGGTCGTCAGGAGATAACTCAGAAGATTTGATGCTGGACATTCACAA
CACGTCGTCGTCTTCGTCTAGACGACCGCCGCCTGAACCATGCTCGAtgatacaacaacaacaacaacaaggaatA
AGAGCACCATCGTCAGCTCTGTGGAGCAATTCTTCGAGGTACGTATACGTAAGCATGCATTTTGTAATTTGCTTTA
ATTTCCTAGCTAGCTTGAAAAGAATGGACAAAAGAGAGATATGGGGGTAGTGTTACATCTTTTTGGCATTATATCT
TTGCATAAAGCACCAACAtgcatttcttttgttaatatactactactactactactactactgttaCTTCTCATTG
TTACGTTATTCCCTCTCCTTTTTCCCTTTAATTCCTACTTTCCCAAATGCACAACTTTTGTTCCTTATGAAcatcc
aaaaaaatacaaaaggaaacaaagagtAAAGGGGGACTTTGATATACACTTGTACCACTGTTAAGTTGCATGTGCA
TGGTGGAAGATGAATAAAGTATAAAAACCTTATATATGCCACTCTCTTTTTCCCAGAGCTTAGGCATGGAAATCCAC
CATCTTTCTGACAAAGGAA
SEQ ID No.8:
GTATTTTATGGGACGACAGTTAGGGGGTACAAGCTGCTAGATCCGTATTTCGTATTCATGAACCCTAT
GCCGACTTACGAGATCACCTTTTTGAACCAGCTGCCGTCGGAGCTGACGTGTAATGGTGGGGGGAAGTCGGCATTT
GAGGTGGCCAATTATATTCAGAAGGTGCTTGGTGGGACCCTTGGGTTTGAGTGTACGAGTCTGACTAGGAAGGATA
AGTATGCCATGATGGCCGGAACCGACGGCTTGATTAAGACCAAACAGGTGATCAAGAAAGACAATATTGAGAAGAG
CTGATCAAAATCGAGGAGGATAAAGAGTTAAATACACCATCgttgtgtttgtgttgtgATTGAGGaactaattaag
aaagaaTGAACATGATCAAAGCTTATTATTTGTGTAGTAttactttctctttttttatccATCGGCAGGTATTGTt
tagaaattgaatttgtccagaaaaaaggaaataaaaaggtaagcattgttttatatatatatatatatatggttgt
atgataaattaattagatgaaATCCATTTGGAGTGATCAGTGACCAATATTAGAATAGTGACAAACGTGACCTATA
ATCCAACAAGAATGTAGTGGAAAGGTTGTACATTTGTTTAAAGTTGGCCTTGACATATCGTCGGtagaggaaaaag
gaaaactagctagctagtaaaTAACAGTGTTGGTTGATAGCATGTTATGTAGCTACCGAAAACTATGGCTTAAAAC
AGTACTTCTATTTAATGCGTTCGACATAAATTGGTGGGAGGTGCTGACGATGGGAACAATACAACTGCCTCCATGT
TGCAAAGAAAATTCCAagacaataatataaaatagtagattgatgaatgaaatgagGCAAAATtaatgtaatgtaa
tgtaaggttttatatatctttactgaaaaagaatgaagattCACGTGATATATATGTAGTCGGACCTAATCTCCCA
AACCTACATGAACAACTTGC
SEQ ID No.9:
CTCTATTCGCACACTTCCAAAGAAAGGTGGTGcactgtattttcgacacaccttttctcgttataaac
tatatttttgtgtgtcgaaatgtattcaaattcgggttttgagtacatgtttggttctcttgcccatCAGGGtcgt
attatggtgtttggtatgtttttgatacatttcaggtgtcatatgacgtttagagataaaccaagacatgacgggg
atGCCgtaacaactcaaagcgttcgacttgggaagaaagcgtcattgtttagtattttgagcataactcatcgctc
cgaagtccaattgatgcaaggtcagcggcgttggaaagataattcgattgtctacaacttttgtgtgacactatgg
tcaaacCCCCCGCGGAGACCCTCGTGACCCCTGTGGACgacatatacttcattatacacatatttttcgagctctt
ggagagagctagagttcctGTTCActcattctacacatattttcgagctttgagagagagagagagagagagatag
agagagagttcCATAGACcaattgaagcgagagaacatcatcGATTGGcaaaggagcgaagatcggcgacaac
SEQ ID No.10:
tctcttttccaatttcatttctcggGATCCGCCATCTCTCACCTTCCCTTCCTCATATGGCTGCCACC
AcaacctcctcctctgccGGTTTCTTCAACTTCCGCTCCAACTCCCTCAAAGTCCGCCACCCTCTCTCCTCCCCCG
CTGGTTGCAGCAAGCTCGACGGCGTCGCCATGTGGGTCGTCAACGGTGTGGCCGCTGCCTTCTTTGCCTCCTTAGA
CAGATGCTCCTGCATCAGGATCGCCAccgttgatgatgttgaagaaggtAATGACGACGCTCCCTTGATTATGAAC
GACGGCAACGTCGACACCCCACCGTGCCGTGGTGGAGGTggcagcagcaggaggaggaggacggcCACCGCCGGTG
GGAAAGGCAAGAAGGCTACTAATTTGTTTGATGGAGAATCGTCTAATTGATGTTGTATTCCACCACTTACTATGAA
GGAGGTAATCAACAGAATTTTCATCCCTCATCTCTGTCGACTGtacaagttttattattattattattattattat
ttttgttatctaAATTGTTTATCAACGCGGTTTTAAGGTTAATTGATGAACACTAATTATATCATCATcgtatttt
ttctttctttctgcgGATGATCAAAGTATATGCCATATATATCCTGAGTTTCATTTTCAGTGAAGAATCGGAGTAC
AATTCCATCCCCAACCATTTTTATTAGCTTAAATAAAGTggtaaaatgttttaaaactTTATTTATAGTAGTGTAT
GGGTTACGAAATTTTATTTACAGTAGAGTAGAGTGAATATCCGATAGTTTGTCAGTTAAACAAGGTGAAATAGCCT
GGGAGGGAGGGAAAGGAAGTAGGGAAAATAAAACTCGATCGATGATTCAGAGGCTAAAGTGTCTGACATATCCAgg
gtataaaaaaaataacaaaattaccACGTAATTGCACACGTctttatctatatatatatatattagtatttGAATT
ACCTTCATTGCAACTGCAGG
SEQ ID No.11:
TCACCAAGATTAAATTGATTAACATATTATCGAAAAAGgttttacaaaatttcaacaaaaaccAACCT
TTCCGAAGGGTGAAACCAGTTGTTGTAGAGCAGTTATTCGGTCGCCCAACTTCTCTTTTCGAACCTGAATTTACCA
CAAAAAAGTCGAGATTTTATTGCAAACGCAACCAAAAGCtctaaaaaaatacaagaacTCGAATTCCAAAAATTCG
ATTTCATTACCACGAAAAATAGTTATTTCATGTgtgtgatgatgatcatctcGAGATTTGTAAACTCCACTATTGA
GTTATGATCACAACATATGATGATTACCTTAAAAGTTGGCAACGAGGATGGAGTCTCGATTCGAGGTCGTTTGAAA
GACGACTCGGAACCggtttccttctttgttttcgGTTTCGACTCCTTGGTAGTATTATTAGtagcttcttccttct
tagACTGTCGAAagtaatcaataaaaaatgtaaaatacaagttaagaaaaaatatatatatatatatataatatat
atatgtcatttGAAACCCTAATGGAGAAGTCATTTTCTTCATACCTTGGTGGAGCTGCTACTGGGGCAACTGGTTG
GTTTCTCGTCAAAAGCCGGGGCAACGAACCGGGTTTGTGTCGGTGCATAACCGGTTCGAATACTATCGGTCTGGGT
TGCTGCTTGATTGTTCCAAAACGGCATCACACCATTCGAGAACCGAAGGTGATCTTGGTGGTGATTGTCGTTTtga
tgattagggttttggaattTCGACCACGAAGGGGACAATCCATTCAACAACAAACTGTTTCCGGTCCCGTAAGTAG
TAGAAGAGGACGGGAAATTGTTGACGAATATCGGGTTTTGTTGCGAATCGGATTCGAACAGGCCTTGAATCGGGTAG
CCATGAATCGGATTCGAAGAGAAATCATTGGTCAACCCACTTAGATTATGGTTGAGTGGACGACCGCAGCTGTTCAT
TTCATCTTCCAGCATTGA
The primer sequence of SSR molecular marker:
RM4-5-F:GTCTCGATCTTTGCATACAGAAGC (as shown in SEQ ID No.12)
RM4-5-R:GTCAAAACATAGCACAACCCCATT (as shown in SEQ ID No.13)
RM4-7-F:GTGATCTTGTTCTTGCTTCACCTC (as shown in SEQ ID No.14)
RM4-7-R:CCCATCGACTTCCCCTAATAGTTT (as shown in SEQ ID No.15)
RM6-2-F:TAGGGTTAGGGTTTCGAATTCCAC (as shown in SEQ ID No.16)
RM6-2-R:ATACGATACGAGACGCTACACAC (as shown in SEQ ID No.17)
RM6-6-F:TCCCTTTTCATCTCTCTCCTCTCT (as shown in SEQ ID No.18)
RM6-6-R:TTCAATGGAAAGGAAATCGATGGC (as shown in SEQ ID No.19)
RM6-7-F:TCTTGCGTTAGAAGATGGAATTGC (as shown in SEQ ID No.20)
RM6-7-R:CAAGAATGAAGGAAGGGGAGAAGA (as shown in SEQ ID No.21)
RM7-1-F:CGTAATGAAGTGGATCTCTTTGGC (as shown in SEQ ID No.22)
RM7-1-R:TCTTCCGCCTTCCTATCATTTTCA (as shown in SEQ ID No.23)
RM7-3-F:ACTCAGAAGATTTGATGCTGGACA (as shown in SEQ ID No.24)
RM7-3-R:CTGACGATGGTGCTCTTATTCCT (as shown in SEQ ID No.25)
RM8-1-F:TCCATCGGCAGGTATTGTTTAGAA (as shown in SEQ ID No.26)
RM8-1-R:TGGATTATAGGTCACGTTTGTCACT (as shown in SEQ ID No.27)
RM9-4-F:GAGAGAGCTAGAGTTCCTGTTCAC (as shown in SEQ ID No.28)
RM9-4-R:TGTTCTCTCGCTTCAATTGGTCTA (as shown in SEQ ID No.29)
RM13-5-F:TTTTCATCCCTCATCTCTGTCGAC (as shown in SEQ ID No.30)
RM13-5-R:GGCATATACTTTGATCATCCGCAG (as shown in SEQ ID No.31)
RM15-1-F:AGCTTCTTCCTTCTTAGACTGTCG (as shown in SEQ ID No.32)
RM15-1-R:CTTTTGACGAGAAACCAACCAGTT (as shown in SEQ ID No.33).
DNA is extracted: extracting 26 parts of flax germplasm, 100mlTPS formula are as follows: 10 milliliters of 1MTris-Hcl (PH using TPS method
=8.0), the EDTA (PH=8.0) of 2 milliliters of 0.5M, 7.45 grams of Kcl are mended with water to 100 milliliters.
SSR primer amplification and electrophoresis detection: 10 μ l reaction systems include: 50 μM of dNTPs, 0.2 μM of primer, 0.5U Taq
Polymerase (TaKaRa, Dalian) and 30ng of DNA profiling.Amplified reaction carries out on LongGeneA200PCR instrument: 95
℃5min；94 DEG C of 30s, (Tm value) 1min, 72 DEG C of 40s, 35 circulations；72 DEG C of extensions 5min, 10 DEG C of preservation 10min.Amplification produces
Object is separated with 8% Native PAGE glue, is developed the color by silver staining, silver staining program is according to Sanguinetti et al.'s (1994)
Method is formulated.The lamp box that the DNA band of amplification is equipped with fluorescent lamp is observed.
Using Flax Varietieies " CDC Bethune " DNA as the PCR product clip size of template amplification: RM4-5 166bp,
RM4-7 is 194bp, and RM6-2 189bp, RM6-6 150bp, RM6-7 172bp, RM7-1 183bp, RM7-3 are
123bp, RM8-1 176bp, RM9-4 112bp, RM13-5 169bp, RM15-1 181bp.
Beneficial effects of the present invention: SSR molecular marker of the present invention is reproducible, band is clear, rich polymorphism
And chromosome physical location information is clear, and has successfully carried out analysis of genetic diversity to 26 parts of Flax Germplasms.This
Invention can be applied to the Genetic diversity evaluation of Flax Germplasm, and Germplasm Identification, the positioning of QTL/ gene and molecular labeling are auxiliary
Help breeding research.
Raw materials used and reagent is available on the market in flax SSR molecular marker provided by the invention and its application.
Below with reference to embodiment, the present invention is further explained:
The acquisition in 1 site flax full-length genome SSR of embodiment
NCBI first downloads flax whole genome sequence information
(https://www.ncbi.nlm.nih.gov/assembly/GCA_000224295.2), further utilize
MISA software (MIcroSAtellite identification tool)
(http://pgrc.ipk-gatersleben.de/misa/) detect flax genome sequence in microsatellite and
Compound microsatellite locus.The detection parameters used is 2-6,3-5,4-5,5-5, the 6-5 (repetition time of 2 base repetitive unit motif
Number >=6 times；The number of repetition of 3 base repetitive units >=5；And so on.)；The distance between two microsatellites are less than 100bp
When, then a compound microsatellite is formed, 28571 sites SSR are detected in entire flax genome.
2 flax SSR design of primers of embodiment and synthesis
According to the SSR testing result of MISA, Primer3 is utilized
(http://pgrc.ipk-gatersleben.de/misa/primer3.html) software default parameter (length
100bp-300bp；55 DEG C -60 DEG C of Tm value, primer length 20-25bp) design of primers is carried out, primer dimer is avoided as far as possible
(dimer), hairpin structure (hairp-in), mispairing (false primer) etc..Highest 3 pairs of marking are selected in design result
Primer is used for subsequent experiment, and 71184 pairs of SSR primers are obtained.Then 96 pairs of primers are therefrom selected at random, it is raw by the Hunan section of holding up
The synthesis of object company.
The amplification of embodiment 3SSR primer and the analysis of genetic diversity of 26 parts of Flax Germplasms
26 parts of flax germplasm (being shown in Table 1) are extracted using TPS method, specific method refers to (Zhang Youchang etc., 2016).Then sharp
Every part of germplasm DNA concentration is measured with DNA, is finally diluted to working solution 30ng/ μ L.SSR reaction system is 10 μ L comprising 50 μM
DNTPs, 0.2 μM of primer, 0.5U Taq polymerase (TaKaRa, Dalian) and 30ng of DNA template.PCR reaction exists
It is carried out in LongGeneA200 gene-amplificative instrament.Reaction total system includes: that PCR reaction is poly- using the rTaq DNA of Takara company
Synthase carries out, and reaction system is following (10 μ L): DNA profiling (10ng/ μ L) 2.0 μ L, primer (2pmol/ μ L) 1 μ L, 10 ×
1.0 0.3 0.1 μ L, ddH2O5.4 μ L of μ L, rTaq (5U/ μ L) of μ L, dNTP (1mM) of Buffer.PCR amplification program includes: specific
Program is as follows: 95 DEG C of initial denaturation 5min；95 DEG C of denaturation 30s, 50-60 DEG C of annealing 30s, 72 DEG C of extension 40s altogether follow for 35 times
Ring；72 DEG C of extension 5min, 4 DEG C of constant temperature save.
PCR product is detected using 8% native polyacrylamide gel electrophoresis.It is added into the PCR product expanded
Two-colour indicator comprising bromophenol blue and dimethylbenzene blueness clicks and enters each 2 ul of PCR sample in electrophoresis tank sample hole, while 500bp
DNA Ladder be control point into loading wells, electrophoresis is finally carried out with 200-230V voltage, the time is with PCR fragment size
Foundation, about 1h.Polyacrylamide gel colour developing: the polyacrylamide gel after electrophoresis is put into 0.1%AgNO3 solution,
12min is at the uniform velocity shaken in shaking table, the colour developing that 1.5%NaOH and 0.4% formaldehyde is added afterwards twice is then cleaned with 400mL ddH2O
Liquid, be placed on shaking table concussion colour developing until band it is clear, finally cleaned 3-5 times with tap water, after preservative film is encapsulated on viewbox
Read recording mark genotype data.It is as follows that genotype data reads standard: the same primer pair difference germplasm DNA is expanded
Increasing, secondly it is B that maximum band, which is denoted as A, and so on, band is unintelligible or missing is denoted as 0.Utilize PowerMarker
3.25 calculate the number (being shown in Table 2) of each primer PIC value and allele.The genetic distance of germplasm is based further on to flax kind
Matter resource carries out clustering.
Table 1
Table 2
| SSR marker | Number of alleles | Genetic distance | PIC value |
| RM4-5 | 2.0000 | 0.5000 | 0.3750 |
| RM4-7 | 4.0000 | 0.5185 | 0.4193 |
| RM6-2 | 3.0000 | 0.4652 | 0.3735 |
| RM6-6 | 3.0000 | 0.4652 | 0.3735 |
| RM6-7 | 3.0000 | 0.5947 | 0.5080 |
| RM7-1 | 4.0000 | 0.5185 | 0.4193 |
| RM7-3 | 4.0000 | 0.6040 | 0.5390 |
| RM8-1 | 6.0000 | 0.7382 | 0.6970 |
| RM9-4 | 3.0000 | 0.4889 | 0.4013 |
| RM13-5 | 2.0000 | 0.4712 | 0.3602 |
| RM15-1 | 5.0000 | 0.7550 | 0.7133 |
By can be seen that in Fig. 1-12,11 sites SSR of the invention are shown preferably in 26 parts of flax germplasm for examination
Polymorphism amplification, and 26 parts of germplasm are preferably divided into two Groups, and Group1 is flax wild species, and Group2 is ground
The kind of square kind and artificially breeding.Group2 is Subgroup1 and Subgroup2, Subgroup1 by further cluster simultaneously
Based on oily kind, Subgroup2 is based on fine kind.The spy of 26 parts of flax germplasm shown in this test result and table 1
Property is coincide substantially.It can be seen that this 11 SSR markers of the invention have reproducible, polymorphism height, the good spy of accuracy
Point is a kind of reliable and effective molecular labeling, can be used for flax analysis of genetic diversity, the building of genetic map and gene/QTL
Positioning.
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Sequence table
<110>Hemp Inst., China Academy of Agricultural Sciences
<120>flax SSR molecular marker and its application
<130> MP1900015
<160> 33
<170> SIPOSequenceListing 1.0
<210> 1
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 1
ttggtcgtgt gtacggagtt aatatggaaa atacacgcag agatttggat acgattttag 60
gtttctacct tcctatctca tactcacgtg tcgttacaac aatgtcattt ggtatagtgc 120
taactaaaaa gttcaatttt tttattataa atagctcaca aatctggacg atcttaaaga 180
caggactatc gactccatag caatttcata tcgaaagacg gaggatgaga taggaccaaa 240
gtcatactaa cctacaaaat cccaatttcc atttttgctc ttttactgag atatgtacac 300
ttgaatttca attttgacat agtttaaaca gtaatttttc aaacttttaa agggaaaaca 360
taaattttac aaacttttaa aggaaaaagt aatgggaggt aaagttgaaa ggaacacagg 420
aattgatgta tgtgaaaatt tgccatgata acagggaaca ttggtgcatg aagggtctcg 480
atctttgcat acagaagcaa ataataataa taataataat aataataatc tcttctcatg 540
tgacattggg aataataatc gtaagattta atacgagtat gtctctttat ttacagatct 600
acactatcat taccccaatg gggttgtgct atgttttgac ctatgaacca tctttcatct 660
cgtagcggcg atagaagtga gaaaaacgta attaaaaatg ttagattaat aacgagagtg 720
tgtgaattac aaaatgtgaa aggacagtct catttaaatt atcatgtaaa gattattagg 780
aatcgtcgaa aaatcaaagt ttaaactatt gaatgtctaa aatgacaaaa tgatgcacat 840
ttacgcgatt ttggaaggct cgatgatgct acaacatctc gcaagctaat ctcctagtga 900
ttctcttatg catctatata ccacaacgaa acttatcatg ttgtctttca ttaaccacta 960
atttttttat gtgactactt acttactagg ttggggatgg 1000
<210> 2
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 2
aaggtcagtc ccacgttcat ctccttggct gcctccgtta tcggcatgta gaaatacctc 60
gacagcattt ccctcgtgat cacctgctgc ccgtttggcg aaaccgcttt gttatgctca 120
ttcgttgttg ttttctgttc atgatcaagg attgggagga gaggtggccc cagttcgttg 180
caaaccggat cgtgatcgat gtgggaagaa catgaagatg atgatattga tgaagatgac 240
gaatgatgaa gaatattagt actactgatc atcatattca tgatgtcagt accgtagcga 300
tcaccatcgg cagcattaac tatatcggga tcttggtagt attgatcaag ctgggtaaaa 360
ggtgcttcga gttcgaactg atgattgaat aatgggaatg gaggaaggga tgatgatgag 420
tagtagtcgt catccccacc accatggctg ctgtacagac acatatatag aaagagggaa 480
tattattaat tatgatggcg atgatgatga tgatgtgtaa gtaattaatt aacgtacgat 540
attacccaaa agtgaagtcg tcaaagagtc ctccgaatgg cgcttgttgc tgggaatcgt 600
catggagaag tggcggtgat gaacggattt ggaagtgatc ttgttcttgc ttcacctcaa 660
caaactccat tgtttgatga tgatgatgat gacgatgaga acacagagcc aaatgaaggg 720
agggggagtg gtttatatgt agagtgcggg tgatgaatga tgattgtgtg cagtaaaaaa 780
tctagtggat taaaaaagct gagaaaacta ttaggggaag tcgatgggtt gctgttttgg 840
aggagatgca taattacagt gagagacgga aaggggagtt acttttttta catattggat 900
tagtgatagt gattaataat taatatcgga gtagtaattg cagtaattca ttgtaatttt 960
aatgggggga attcattatt gatgtcaata taattaacac 1000
<210> 3
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 3
cacttggtcg atagcgtcct ccgatactct gcctccgcca atccggacgg agccggaaga 60
aagaccagct ggagccgaaa ccaaatcatc aggtagttcc agctgagtcg attgtagctt 120
ggagtccttg agctttaaac gattcatagt atcgtccaaa tcggccaact cccatgaatc 180
cggggctccg agatcatctg acacagagcc ctccatggat tctttgacgc agagagctaa 240
cggaataagg aggtgaacgc taagtcacct gaaaaggaac cttaaagtgc aaatcaatcg 300
tcgctagggt tgtcgacgtt ccagcaatat tgtgttgagc atcgtgttat atgaccgtaa 360
atcgaccgat ccaggattgc gattaggccg acgactccga ggcgtgaatc acatgcagaa 420
tgcgactcgt agggttaggg tttcgaattc caccggatta tttggctaaa attggagagg 480
cctacaatct ttcttacata gagagagaga gagagagaga gagggagaaa gagagagaga 540
gagaatgaga gaggatagag tttgataagt cgtttgtcga aattcgaaag ggggtgtgtg 600
tagcgtctcg tatcgtatgc gcttaagcac agtagaaaaa acgaaaggtc ggaaatttaa 660
aatataagag atttaaataa attttattat ttttaaaata atttataaga aattttaaat 720
atgattatta gtgtagtaat ctattgataa gttgataatt tgattagtaa taattgtata 780
ataagccaat ttagtttagc agaatgcgaa ttttattaga aataaattaa atagtgtttt 840
atttgtttaa atgtagttgt tgaaaaaaat ggcgaagaca aagggtggag ctaaatcgta 900
caattaagat ccggtagagg aagaaacgaa tggaattcag aaaatgagag aaaaaggaaa 960
gcaagctggc attttaaatc gaatgaaaga gcgagtgatc 1000
<210> 4
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 4
aaacagtgac gctttcttcc aaagtcgaac gctttgagtt gttccggtct cctcatcatg 60
ttttggttta tctccaaacg tcatatgaca cctgaaatat accaaaaaca taccaaacac 120
cataataccg ccccgagggg caagagaacc aaacatgtac tcacaaccca aatttgaata 180
catttcgaca cacaaaaata tagtttataa cgagaaaagg tatgtcgaaa atacatcaca 240
tcactgctaa ctacaataac acttaattag taatgaccac aacttgattt ttcacctttt 300
tttcaaaagg caaatacttc aactttgata aaaatgaatc aaaattcagc ctctcgacta 360
ataacatcag aaatctcaaa agccattaaa tttcaatgag caagtctatt tgcttctcga 420
cagacaaaac tccaatctca aatatcacaa cgagcaattc cactcgacag acgctgccag 480
cttcagtcct gcaaatatcc ttcttcttct tcttcttctt cttcagcaaa tcaataagct 540
ttttacagtc caatttttga acttaaaaat gggggagagt gtgttcaaaa ttaaattaaa 600
gacaaaaagc ctcaacataa ttcttgttgt ttgagtgaga gtgcgtgtgg ttttcggagt 660
gtttgcatga tatgatataa tatgatatga ccccatccat cttcttcctc cctcaatatc 720
atataaaatc aaagtttctt tctttctttc cccttttccc ttttcatctc tctcctctct 780
ctctatctct ctgtactctg tgtctctttc tttttcagct gcttcttctt cttcttcttc 840
ttcttcttcc tggtgatggc tttttcatct gtacctctgt cggccatcga tttcctttcc 900
attgaatctt tgatatccac atgaaagaag attccttcgc tgcctacttt ttttctcttg 960
attttccttc ttctcccccc taaggtcatg tgattcaggc 1000
<210> 5
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 5
ttccagctgt cccacgtgtc gtgcatccat gatcccagta ggtcctttgt cgacgtgtgc 60
cagacgggct gacgtggaag cttacagtgt acgaggcaat atggttggag atgaatttgg 120
agttgctcga taatgtaaaa ttgattcgac gatataatta taattttatg aacaattggt 180
gcattattaa gattcctctg tgtcctctat ttccccccca tattttaacc cagagatgga 240
tggggagaat tatgtgcaag aaatgaaggg tgaagaaaag aatatgatgt gttgagcatg 300
tgaatataca aagtcatgta aatagaaaga cgattcctgt ttaggaagca aagaaaggtg 360
ctcgacgggg aatccctttt cttcttcttg cgttagaaga tggaattgct ttgtacatat 420
cttctccttg ttgcaattca tgttcatttg gtgatgacga tgacgacatc atcgctctag 480
acctccctat aactcttctt tcttcttctt cttcttcttc ctcgtttgct tcttcttctc 540
cccttccttc attcttgcct tccttgctaa accttgcatt cccattcttc tttattccaa 600
caagctacaa ggacataacc atgtaattaa ttccccacta attaacttgt tatcacattt 660
ggtcaaaacc gtattatatt accactcggc caattccttg gagttctgaa ttcggacact 720
ttagtaactt ataccgtaaa gtgtcattag gtaaattatt tttcaatcca acacatgcat 780
tgataatctt gatttccata gtcaactttt tgaaagatgg aaaaccaacc ataggtgatg 840
atgatgaagc ttgacgtatt aaaaacatca tttgggttag gtattttata tatgtagtat 900
tttgaaacat tatttaaaga tatggtattt atgagataaa atttgtgatg tgtaaattat 960
cttgtttaac tattcaacgt ggtagtttat cggggtattt 1000
<210> 6
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 6
tacaaattcg gtgtgggtcg ataatggaca gctgactcat atctagtcga attagtccaa 60
gcaggtcgag tttgaattgt catttgagag gaagagaatc aaatggagga ttcatcaata 120
tacaccactt ctttacattg aataacaaac tcaaactctt ggggcacaaa aaaatttggg 180
aagagatcga gttcattcat ctctttgaac tcaatataaa gactgttcga atcttgagac 240
caacaatcat ctattatcta acaggcatag ggagcgaaaa ccctaacagc tgagcgcgga 300
ggtgagacta acgaagtact cctcgctgca cgggatggtg agtccgccca tggggtgatc 360
gaacccgaac tcttcctcgg ctcgatcgag taagacttgg aacaaagggt ggttcaagta 420
agtgactggg atgatgaacc tcttcttctt aaggtagcct tcaccaacat agactgcaat 480
atggcccttt ggcacattgg aagaagaaga agaagaagat agtttggctg acagtgttct 540
ttgtagcttt tgccttgcat tttcagcaat ccccattgtt aagaaacaga ggttgagaaa 600
cagaggaaaa gaaggagttt ttattggtgg gttttgagtt tttactaagt ttgtttcatg 660
catgaaggct tatatataga gagaggtggg gttttgcaag tgaaattctt gaaagaataa 720
ttgtaagtag aagaagaaga agaaacatac caggtgaaga ggctgctgaa gattcacatg 780
ggtgtgcttg tgtccatggc tacatcctct ggcctactgt ctttttcagt tgagtgctat 840
gagctacata ttctgtgttc acataagagt tcaaagtttg cttatctaag caaggcaagg 900
ggtttaacta cgttcaattt cgttattcag ccacacccta accgacagtg ccctcaactg 960
agtggcttta ggcacaacgt gttgcataga agtagcttga 1000
<210> 7
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 7
aggcgggttg atcaggttaa catttatagg ggttcaatca tgtgacattt gtttgtattc 60
gggttggggt tgggttgggg ggggaggtta ttagattaat ggcttctagt cgattcaaat 120
cattaaccga gtcagataag actcgaactg agtggatgca agatcagctt cttaattaca 180
attcttattg tataatccac attaactgat taacggtggt tggttgattg cagatgtacc 240
gtacggtaaa gaccactgac agagctgctg ccacgtcagg ttaacgactt aacccccaaa 300
atgaacttac ttatatgatc aatatcaatg gttgcttcat gatatttacg gaaatacttt 360
tgcaacatat gtttacaggg caatcagatt tagttgacaa cgggtcgtca ggagataact 420
cagaagattt gatgctggac attcacaaca cgtcgtcgtc ttcgtctaga cgaccgccgc 480
ctgaaccatg ctcgatgata caacaacaac aacaacaagg aataagagca ccatcgtcag 540
ctctgtggag caattcttcg aggtacgtat acgtaagcat gcattttgta atttgcttta 600
atttcctagc tagcttgaaa agaatggaca aaagagagat atgggggtag tgttacatct 660
ttttggcatt atatctttgc ataaagcacc aacatgcatt tcttttgtta atatactact 720
actactacta ctactactgt tacttctcat tgttacgtta ttccctctcc tttttccctt 780
taattcctac tttcccaaat gcacaacttt tgttccttat gaacatccaa aaaaatacaa 840
aaggaaacaa agagtaaagg gggactttga tatacacttg taccactgtt aagttgcatg 900
tgcatggtgg aagatgaata aagtataaaa accttatata tgccactctc tttttcccag 960
agcttaggca tggaaatcca ccatctttct gacaaaggaa 1000
<210> 8
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 8
gtattttatg ggacgacagt tagggggtac aagctgctag atccgtattt cgtattcatg 60
aaccctatgc cgacttacga gatcaccttt ttgaaccagc tgccgtcgga gctgacgtgt 120
aatggtgggg ggaagtcggc atttgaggtg gccaattata ttcagaaggt gcttggtggg 180
acccttgggt ttgagtgtac gagtctgact aggaaggata agtatgccat gatggccgga 240
accgacggct tgattaagac caaacaggtg atcaagaaag acaatattga gaagagctga 300
tcaaaatcga ggaggataaa gagttaaata caccatcgtt gtgtttgtgt tgtgattgag 360
gaactaatta agaaagaatg aacatgatca aagcttatta tttgtgtagt attactttct 420
ctttttttat ccatcggcag gtattgttta gaaattgaat ttgtccagaa aaaaggaaat 480
aaaaaggtaa gcattgtttt atatatatat atatatatgg ttgtatgata aattaattag 540
atgaaatcca tttggagtga tcagtgacca atattagaat agtgacaaac gtgacctata 600
atccaacaag aatgtagtgg aaaggttgta catttgttta aagttggcct tgacatatcg 660
tcggtagagg aaaaaggaaa actagctagc tagtaaataa cagtgttggt tgatagcatg 720
ttatgtagct accgaaaact atggcttaaa acagtacttc tatttaatgc gttcgacata 780
aattggtggg aggtgctgac gatgggaaca atacaactgc ctccatgttg caaagaaaat 840
tccaagacaa taatataaaa tagtagattg atgaatgaaa tgaggcaaaa ttaatgtaat 900
gtaatgtaag gttttatata tctttactga aaaagaatga agattcacgt gatatatatg 960
tagtcggacc taatctccca aacctacatg aacaacttgc 1000
<210> 9
<211> 597
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 9
ctctattcgc acacttccaa agaaaggtgg tgcactgtat tttcgacaca ccttttctcg 60
ttataaacta tatttttgtg tgtcgaaatg tattcaaatt cgggttttga gtacatgttt 120
ggttctcttg cccatcaggg tcgtattatg gtgtttggta tgtttttgat acatttcagg 180
tgtcatatga cgtttagaga taaaccaaga catgacgggg atgccgtaac aactcaaagc 240
gttcgacttg ggaagaaagc gtcattgttt agtattttga gcataactca tcgctccgaa 300
gtccaattga tgcaaggtca gcggcgttgg aaagataatt cgattgtcta caacttttgt 360
gtgacactat ggtcaaaccc cccgcggaga ccctcgtgac ccctgtggac gacatatact 420
tcattataca catatttttc gagctcttgg agagagctag agttcctgtt cactcattct 480
acacatattt tcgagctttg agagagagag agagagagag atagagagag agttccatag 540
accaattgaa gcgagagaac atcatcgatt ggcaaaggag cgaagatcgg cgacaac 597
<210> 10
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 10
tctcttttcc aatttcattt ctcgggatcc gccatctctc accttccctt cctcatatgg 60
ctgccaccac aacctcctcc tctgccggtt tcttcaactt ccgctccaac tccctcaaag 120
tccgccaccc tctctcctcc cccgctggtt gcagcaagct cgacggcgtc gccatgtggg 180
tcgtcaacgg tgtggccgct gccttctttg cctccttaga cagatgctcc tgcatcagga 240
tcgccaccgt tgatgatgtt gaagaaggta atgacgacgc tcccttgatt atgaacgacg 300
gcaacgtcga caccccaccg tgccgtggtg gaggtggcag cagcaggagg aggaggacgg 360
ccaccgccgg tgggaaaggc aagaaggcta ctaatttgtt tgatggagaa tcgtctaatt 420
gatgttgtat tccaccactt actatgaagg aggtaatcaa cagaattttc atccctcatc 480
tctgtcgact gtacaagttt tattattatt attattatta ttatttttgt tatctaaatt 540
gtttatcaac gcggttttaa ggttaattga tgaacactaa ttatatcatc atcgtatttt 600
ttctttcttt ctgcggatga tcaaagtata tgccatatat atcctgagtt tcattttcag 660
tgaagaatcg gagtacaatt ccatccccaa ccatttttat tagcttaaat aaagtggtaa 720
aatgttttaa aactttattt atagtagtgt atgggttacg aaattttatt tacagtagag 780
tagagtgaat atccgatagt ttgtcagtta aacaaggtga aatagcctgg gagggaggga 840
aaggaagtag ggaaaataaa actcgatcga tgattcagag gctaaagtgt ctgacatatc 900
cagggtataa aaaaaataac aaaattacca cgtaattgca cacgtcttta tctatatata 960
tatatattag tatttgaatt accttcattg caactgcagg 1000
<210> 11
<211> 1000
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 11
tcaccaagat taaattgatt aacatattat cgaaaaaggt tttacaaaat ttcaacaaaa 60
accaaccttt ccgaagggtg aaaccagttg ttgtagagca gttattcggt cgcccaactt 120
ctcttttcga acctgaattt accacaaaaa agtcgagatt ttattgcaaa cgcaaccaaa 180
agctctaaaa aaatacaaga actcgaattc caaaaattcg atttcattac cacgaaaaat 240
agttatttca tgtgtgtgat gatgatcatc tcgagatttg taaactccac tattgagtta 300
tgatcacaac atatgatgat taccttaaaa gttggcaacg aggatggagt ctcgattcga 360
ggtcgtttga aagacgactc ggaaccggtt tccttctttg ttttcggttt cgactccttg 420
gtagtattat tagtagcttc ttccttctta gactgtcgaa agtaatcaat aaaaaatgta 480
aaatacaagt taagaaaaaa tatatatata tatatataat atatatatgt catttgaaac 540
cctaatggag aagtcatttt cttcatacct tggtggagct gctactgggg caactggttg 600
gtttctcgtc aaaagccggg gcaacgaacc gggtttgtgt cggtgcataa ccggttcgaa 660
tactatcggt ctgggttgct gcttgattgt tccaaaacgg catcacacca ttcgagaacc 720
gaaggtgatc ttggtggtga ttgtcgtttt gatgattagg gttttggaat ttcgaccacg 780
aaggggacaa tccattcaac aacaaactgt ttccggtccc gtaagtagta gaagaggacg 840
ggaaattgtt gacgaatatc gggttttgtt gcgaatcgga ttcgaacagg ccttgaatcg 900
ggtagccatg aatcggattc gaagagaaat cattggtcaa cccacttaga ttatggttga 960
gtggacgacc gcagctgttc atttcatctt ccagcattga 1000
<210> 12
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 12
gtctcgatct ttgcatacag aagc 24
<210> 13
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 13
gtcaaaacat agcacaaccc catt 24
<210> 14
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 14
gtgatcttgt tcttgcttca cctc 24
<210> 15
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 15
cccatcgact tcccctaata gttt 24
<210> 16
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 16
tagggttagg gtttcgaatt ccac 24
<210> 17
<211> 23
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 17
atacgatacg agacgctaca cac 23
<210> 18
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 18
tcccttttca tctctctcct ctct 24
<210> 19
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 19
ttcaatggaa aggaaatcga tggc 24
<210> 20
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 20
tcttgcgtta gaagatggaa ttgc 24
<210> 21
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 21
caagaatgaa ggaaggggag aaga 24
<210> 22
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 22
cgtaatgaag tggatctctt tggc 24
<210> 23
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 23
tcttccgcct tcctatcatt ttca 24
<210> 24
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 24
actcagaaga tttgatgctg gaca 24
<210> 25
<211> 23
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 25
ctgacgatgg tgctcttatt cct 23
<210> 26
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 26
tccatcggca ggtattgttt agaa 24
<210> 27
<211> 25
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 27
tggattatag gtcacgtttg tcact 25
<210> 28
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 28
gagagagcta gagttcctgt tcac 24
<210> 29
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 29
tgttctctcg cttcaattgg tcta 24
<210> 30
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 30
ttttcatccc tcatctctgt cgac 24
<210> 31
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 31
ggcatatact ttgatcatcc gcag 24
<210> 32
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 32
agcttcttcc ttcttagact gtcg 24
<210> 33
<211> 24
<212> DNA
<213>artificial sequence (Artificial Sequence)
<400> 33
cttttgacga gaaaccaacc agtt 24
Claims (10)
1. flax SSR molecular marker, which is characterized in that flax SSR molecular marker has as appointed in No.1~11 SEQ ID
One or appoint several shown in sequence.
2. flax SSR molecular marker as described in claim 1 is in Flax Germplasm analysis of genetic diversity, flax high density
Application in the building of map, Flax Varietieies identification and/or molecular mark.
3. expanding the primer sets of flax SSR molecular marker as described in claim 1, which is characterized in that appoint including (1)~(11)
One group or several groups of primers:
(1), expand the upstream primer of the sequence as shown in SEQ ID No.1: its sequence is as shown in SEQ ID No.12；
Expand the downstream primer of the sequence as shown in SEQ ID No.1: its sequence is as shown in SEQ ID No.13；
(2), expand the upstream primer of the sequence as shown in SEQ ID No.2: its sequence is as shown in SEQ ID No.14；
Expand the downstream primer of the sequence as shown in SEQ ID No.2: its sequence is as shown in SEQ ID No.15；
(3), expand the upstream primer of the sequence as shown in SEQ ID No.3: its sequence is as shown in SEQ ID No.16；
Expand the downstream primer of the sequence as shown in SEQ ID No.3: its sequence is as shown in SEQ ID No.17；
(4), expand the upstream primer of the sequence as shown in SEQ ID No.4: its sequence is as shown in SEQ ID No.18；
Expand the downstream primer of the sequence as shown in SEQ ID No.4: its sequence is as shown in SEQ ID No.19；
(5), expand the upstream primer of the sequence as shown in SEQ ID No.5: its sequence is as shown in SEQ ID No.20；
Expand the downstream primer of the sequence as shown in SEQ ID No.5: its sequence is as shown in SEQ ID No.21；
(6), expand the upstream primer of the sequence as shown in SEQ ID No.6: its sequence is as shown in SEQ ID No.22；
Expand the downstream primer of the sequence as shown in SEQ ID No.6: its sequence is as shown in SEQ ID No.23；
(7), expand the upstream primer of the sequence as shown in SEQ ID No.7: its sequence is as shown in SEQ ID No.24；
Expand the downstream primer of the sequence as shown in SEQ ID No.7: its sequence is as shown in SEQ ID No.25；
(8), expand the upstream primer of the sequence as shown in SEQ ID No.8: its sequence is as shown in SEQ ID No.26；
Expand the downstream primer of the sequence as shown in SEQ ID No.8: its sequence is as shown in SEQ ID No.27；
(9), expand the upstream primer of the sequence as shown in SEQ ID No.9: its sequence is as shown in SEQ ID No.28；
Expand the downstream primer of the sequence as shown in SEQ ID No.9: its sequence is as shown in SEQ ID No.29；
(10), expand the upstream primer of the sequence as shown in SEQ ID No.10: its sequence is as shown in SEQ ID No.30；
Expand the downstream primer of the sequence as shown in SEQ ID No.10: its sequence is as shown in SEQ ID No.31；
(11), expand the upstream primer of the sequence as shown in SEQ ID No.11: its sequence is as shown in SEQ ID No.32；
Expand the downstream primer of the sequence as shown in SEQ ID No.11: its sequence is as shown in SEQ ID No.33.
4. primer sets as claimed in claim 3 are expanding the application in flax SSR molecular marker as described in claim 1.
5. primer sets as claimed in claim 3 are in Flax Germplasm analysis of genetic diversity, the structure of flax high density map
It builds, the application in Flax Varietieies identification and/or molecular mark.
6. kit, which is characterized in that including primer sets as claimed in claim 3.
7. kit as claimed in claim 6 is in Flax Germplasm analysis of genetic diversity, the structure of flax high density map
It builds, the application in Flax Varietieies identification and/or molecular mark.
8. the method for Flax Germplasm analysis of genetic diversity, which is characterized in that utilize primer sets as claimed in claim 3
Flax sample to be tested is expanded, electrophoresis detection, obtains genotype data；
The genotype data reads standard are as follows: and same group of primer pair difference flax sample to be tested expands, in electrophoresis detection result,
Maximum band is denoted as A, is secondly B, and so on, band is unintelligible or missing is denoted as 0；Utilize PowerMarker 3.25
The PIC value of every group of primer and the number of allele are obtained, Flax Germplasm genetic diversity knot is obtained according to the PIC value
Fruit.
9. the method for Flax Varietieies identification, which is characterized in that using primer sets as claimed in claim 3 to flax sample to be tested
It is expanded, electrophoresis detection, obtains genotype data；
The genotype data reads standard are as follows: and same group of primer pair difference flax sample to be tested expands, in electrophoresis detection result,
Maximum band is denoted as A, is secondly B, and so on, band is unintelligible or missing is denoted as 0；Utilize PowerMarker 3.25
The PIC value of every group of primer and the number of allele are obtained, Flax Varietieies qualification result is obtained.
10. the preparation method of flax germplasm dendrogram, which is characterized in that using primer sets as claimed in claim 3 to flax
Sample to be tested is expanded, electrophoresis detection, obtains genotype data；
The genotype data reads standard are as follows: and same group of primer pair difference flax sample to be tested expands, in electrophoresis detection result,
Maximum band is denoted as A, is secondly B, and so on, band is unintelligible or missing is denoted as 0；Obtain every group of primer heredity away from
From based on the genetic distance to Flax Germplasm progress clustering.
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| CN201910067537.2A CN109517925A (en) | 2019-01-24 | 2019-01-24 | Flax SSR molecular marker and its application |
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| CN201910067537.2A CN109517925A (en) | 2019-01-24 | 2019-01-24 | Flax SSR molecular marker and its application |
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Cited By (7)
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| CN110195126A (en) * | 2019-07-16 | 2019-09-03 | 山西省农业科学院农作物品种资源研究所 | SSR core primers group and its application based on bitter buckwheat full-length genome data mining |
| CN111187854A (en) * | 2020-03-24 | 2020-05-22 | 北京市园林科学研究院 | SSR primer for identifying sedge molecules and application thereof |
| CN112941218A (en) * | 2021-02-04 | 2021-06-11 | 湖北省农业科学院粮食作物研究所 | Method for identifying authenticity of yam germplasm resources by virtue of cpPSSR molecular marker method |
| CN113151536A (en) * | 2021-02-18 | 2021-07-23 | 甘肃省农业科学院作物研究所 | SSR molecular marker detection method for authenticity of conventional variety of oil flax |
| CN113699266A (en) * | 2021-08-27 | 2021-11-26 | 黑龙江省农业科学院农产品质量安全研究所 | Hemp SSR molecular marker and application thereof |
| CN114480696A (en) * | 2020-10-23 | 2022-05-13 | 广西壮族自治区亚热带作物研究所（广西亚热带农产品加工研究所） | AMP deaminase gene SSR molecular marker primer of tea tree and application |
| CN114525345A (en) * | 2022-02-14 | 2022-05-24 | 江苏科技大学 | SSR molecular marker of castor silkworm and application thereof |
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| CN110195126A (en) * | 2019-07-16 | 2019-09-03 | 山西省农业科学院农作物品种资源研究所 | SSR core primers group and its application based on bitter buckwheat full-length genome data mining |
| CN111187854A (en) * | 2020-03-24 | 2020-05-22 | 北京市园林科学研究院 | SSR primer for identifying sedge molecules and application thereof |
| CN114480696A (en) * | 2020-10-23 | 2022-05-13 | 广西壮族自治区亚热带作物研究所（广西亚热带农产品加工研究所） | AMP deaminase gene SSR molecular marker primer of tea tree and application |
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| CN113151536A (en) * | 2021-02-18 | 2021-07-23 | 甘肃省农业科学院作物研究所 | SSR molecular marker detection method for authenticity of conventional variety of oil flax |
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