CN107446991A - A set of SNP site and its application for being applied to identification hop varieties and purity - Google Patents
A set of SNP site and its application for being applied to identification hop varieties and purity Download PDFInfo
- Publication number
- CN107446991A CN107446991A CN201710279239.0A CN201710279239A CN107446991A CN 107446991 A CN107446991 A CN 107446991A CN 201710279239 A CN201710279239 A CN 201710279239A CN 107446991 A CN107446991 A CN 107446991A
- Authority
- CN
- China
- Prior art keywords
- hops
- snp site
- sense primer
- snp
- primer
- 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
- 241000218228 Humulus Species 0.000 title claims abstract 9
- 244000025221 Humulus lupulus Species 0.000 claims abstract description 63
- 235000008694 Humulus lupulus Nutrition 0.000 claims abstract description 49
- 239000012807 PCR reagent Substances 0.000 claims description 24
- 239000002773 nucleotide Substances 0.000 claims description 11
- 125000003729 nucleotide group Chemical group 0.000 claims description 11
- 230000000692 anti-sense effect Effects 0.000 claims description 10
- 238000001514 detection method Methods 0.000 claims description 6
- 238000003205 genotyping method Methods 0.000 claims description 5
- 238000007400 DNA extraction Methods 0.000 claims description 2
- 238000003753 real-time PCR Methods 0.000 claims description 2
- 238000005259 measurement Methods 0.000 claims 1
- 235000013405 beer Nutrition 0.000 abstract description 6
- 238000000034 method Methods 0.000 abstract description 5
- 238000012268 genome sequencing Methods 0.000 abstract description 3
- 238000012216 screening Methods 0.000 abstract description 3
- 238000012795 verification Methods 0.000 abstract 1
- 239000000523 sample Substances 0.000 description 11
- 108020004414 DNA Proteins 0.000 description 7
- 108090000623 proteins and genes Proteins 0.000 description 5
- 238000011529 RT qPCR Methods 0.000 description 4
- 239000002994 raw material Substances 0.000 description 4
- 238000001914 filtration Methods 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000002372 labelling Methods 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- 238000012797 qualification Methods 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 235000018553 tannin Nutrition 0.000 description 2
- 229920001864 tannin Polymers 0.000 description 2
- 239000001648 tannin Substances 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 1
- 241000196324 Embryophyta Species 0.000 description 1
- 241000628997 Flos Species 0.000 description 1
- 206010019133 Hangover Diseases 0.000 description 1
- 108020005120 Plant DNA Proteins 0.000 description 1
- 238000000246 agarose gel electrophoresis Methods 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000000844 anti-bacterial effect Effects 0.000 description 1
- 230000000259 anti-tumor effect Effects 0.000 description 1
- 239000003963 antioxidant agent Substances 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 235000006708 antioxidants Nutrition 0.000 description 1
- 235000019658 bitter taste Nutrition 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- OEYIOHPDSNJKLS-UHFFFAOYSA-N choline Chemical compound C[N+](C)(C)CCO OEYIOHPDSNJKLS-UHFFFAOYSA-N 0.000 description 1
- 229960001231 choline Drugs 0.000 description 1
- 230000001112 coagulating effect Effects 0.000 description 1
- 239000013068 control sample Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 238000000855 fermentation Methods 0.000 description 1
- 230000004151 fermentation Effects 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 229930003935 flavonoid Natural products 0.000 description 1
- 235000017173 flavonoids Nutrition 0.000 description 1
- 150000002215 flavonoids Chemical class 0.000 description 1
- 230000004907 flux Effects 0.000 description 1
- 239000003205 fragrance Substances 0.000 description 1
- 238000000227 grinding Methods 0.000 description 1
- 230000008676 import Effects 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 239000003147 molecular marker Substances 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 238000003752 polymerase chain reaction Methods 0.000 description 1
- 102000054765 polymorphisms of proteins Human genes 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 230000001835 salubrious effect Effects 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 239000000341 volatile oil Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
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
The invention discloses a set of SNP site for being applied to identification hop varieties and purity, altogether including 5 hops SNP sites.The present invention is based on the genome sequencing result to 10 domestic and international representative hop varieties, screening is compared to the SNP site of full-length genome, the identification that preferable 5 SNP sites of polymorphism carry out hop varieties and purity is demonstrated at present, 5 SNP sites can differentiate to hop varieties, and genotypic results are stable, treat completion hops SNP site fingerprint database after the completion of follow-up SNP site authenticity verification work.SNP site provided by the invention and authentication method, a kind of low cost, high efficiency, the hop varieties of standardization and Purity system are provided for Beer Brewage industry.
Description
Technical field
The invention belongs to biological technical field, and in particular to a set of SNP positions for being applied to identification hop varieties and purity
Point and its application.
Background technology
Hops (Humulus lupulus), also known as hops, Flos lupuli flower, hops and cordateleaf European hop flower.Mainly it is distributed in China
In the province such as Xinjiang, northeast, North China and Shandong, Gansu and Shaanxi.Hops is a kind of with valuable economic value and medicinal
The crop of value.Hops contain resinae, volatile oil, flavonoids, tannin, choline, crude fibre, amino acid etc. it is a variety of chemistry into
Point, there is antibacterial, antitumor, anti-oxidant, calm isoreactivity.Resin contained therein assigns the unique fragrance of beer and salubrious
Bitter taste；Tannin can eliminate caused lactic acid bacteria in fermentation process, and can by protein coagulating unnecessary in wine liquid, separate
Clarify wine liquid.Therefore hops is essential primary raw material during beer brewing.In recent years, China's Beer Brewage is looked forward to
Buying of the industry for hop raw material is not only from real estate, and same import hops also accounts for certain market share, and in the market carries
The some hop raw material various degrees supplied mix, and beer enterprise is no to hop raw material simple and effective
Identification of means, economic loss thus is caused to beer enterprise.Therefore, the identification side of the hop varieties of efficiently and accurately and purity
Method needs foundation badly.
DNA molecular marker is the effective means of conventional in recent years identification of species and purity.SNP(single
Nucleotide polymorphism), i.e. SNP is third generation molecular labeling, compared with other molecular labelings
With unrivaled advantage, therefore the mark for being proposed as updating in kind parting context of detection.SNP marker have with
Under advantage：1. hereditary variation is stable；2. it is applied to high flux Large-scale Screening and accurate Genotyping；3. sample DNA is only
Need ng levels.(Laboratory of the Government Chemist, government chemist are real by this research and utilization Britain LGC
Test room) the KASP technologies of Co., Ltd, based on 10 conventional hop varieties genome sequencing results both at home and abroad, excavate
SNP site in hops genome, has filtered out the preferable hops SNP site of 5 polymorphisms, by using conjunction at present
Into 5 groups of KASP specific primers carry out real-time quantitative PCR, according to the fluorescence intensity of PCR primer come to hops carry out kind
And purity detecting.This method establishes standardization, high efficiency, the Molecular Detection platform of low cost, is that hops sample kind is true
Reality and Purity identify good basis.
The content of the invention
1. first purpose of the present invention, it is desirable to provide a set of 5 SNP sites using hops are come to hops progress
Genotyping.The information of 5 described SNP sites is as follows：
SNP site 1 is the Contig LD161349.1 submitted on NCBI the 9970th nucleotides, specific place sequence
It is as follows： GCCACCTACAACAATTCAAGGTGCCAGAAGGACACTTGGATCTCAGCGAACCAGGTCCTGAGGGCGCCCA
CATAAGAATATCAGCCACTCTGTCAGGAAAACAACTCCT(A/C)GTATGCAGTTGCTATTTGGAGGGCAAACA
CCCCAAGTTGAACTACAACGAGTTAATCAGATAGCTCGGGAAAACAGAGACGGACTCCCTTCTAATCCGAC；
SNP site 2 is the Contig LD205009.1 submitted on NCBI the 1776th nucleotides, specific place sequence
It is as follows： CTCGTGGAGAGTGAGACCACGCGGTTCTAAGAACACGCGCCTGTGCTCCCAACAAATCCCCAATTACTCGG
AATTCGTGTGTCAGGAGGGTCAGGTTAGAATTTT(G/T)CCTTGGAGGGAAAGTTCCAATAGGCAAGAAAGG
CAAAACCGCCTGTGAAGCGTGTCCCCTAAGCTACCTGGTTTTGGACCCAAAATACTGGAAA；
SNP site 3 is the Contig LD169680.1 submitted on NCBI the 2676th nucleotides, specific place sequence
It is as follows： GTCAGGAGTTCTCAGATTACAGCTGCAAGAAGAAAGGACAGGTAAGAGAA GAATTGATCTGAAGCCGCAA
AGAAGCTAATGAAGAGCGTCTGAAG(A/G)ATTGAAAGCATAACAAGACTGGTAAGCGCGTCATCTATTGCAC
AAGCACCTCCGGAGACGACTACATCAGAGATGAGAAGAACAGCAAAACTTG；
SNP site 4 is the Contig LD167190.1 submitted on NCBI 16972 nucleotides, and specific place sequence is such as
Under:
TCCCGCAAGGTTCGACCCATCCTCGCGCCAGATGCGCCTGGGCGTGCACAATCCGTGACGCACTCCCACAA
GTCCATGTCCTCATCATAGA(T/C)CACGTCCTTTATGCCACCACTCTTAACTTAACCATAACACACATATATTCC
AGCATCATCAGATGTCAATCAATTCTTACCTTGTCTTCTGAATTTCT；
SNP site 5 is the Contig LD201072.1 submitted on NCBI 2650 nucleotides, and specific place sequence is such as
Under：
GATATTACCAGTTCGCTGACATCTATCACAGGCCTTGACAAAAACATTAGCATCCTTGAATAATGAAGGCCAAT
AGAAACCACTTTGTAATACCTTGGC(T/C)GCCGTTCTTGATGCTCCAAAGTGCCCACCACATTGTTGAGTGTG
ACAATGGTTGAGAATTGACTGCATCTCTTCTTCAGGAACACACCTTCTGATAATCT；
2. second object of the present invention, it is desirable to provide a set of 5 SNP positions for being used to identify hop varieties and purity
Point, the identification of kind and purity is carried out to hops sample with reference to KASP specific primers and PCR reagent.
5 groups of described primers are as follows：
(1) primer sets 1 for detecting hops genome SNP site 1 are：
Sense primer 1：CTGTCAGGAAAACAACTCCTA
Sense primer 2：CTGTCAGGAAAACAACTCCTC
Anti-sense primer：CATACGTCAACGATAAACCT
(2) primer sets 2 for detecting hops genome SNP site 2 are：
Sense primer 1：GAGGGTCAGGTTAGAATTTTG
Sense primer 2：GAGGGTCAGGTTAGAATTTTT
Anti-sense primer：GGAACCTCCCTTTCAAGGTT
(3) primer sets 3 for detecting hops genome SNP site 3 are：
Sense primer 1：CTAATGAAGAGCGTCTGAAGA
Sense primer 2：CTAATGAAGAGCGTCTGAAGG
Anti-sense primer：TAACTTTCGTATTGTTCTGA
(4) primer sets 4 for detecting hops genome SNP site 4 are：
Sense primer 1：GTCCATGTCCTCATCATAGAT
Sense primer 2：GTCCATGTCCTCATCATAGAC
Anti-sense primer：GTGCAGGAAATACGGTGGTG
(5) primer sets 5 for detecting hops genome SNP site 5 are：
Sense primer 1：CCACTTTGTAATACCTTGGCT
Sense primer 2：CCACTTTGTAATACCTTGGCC
Anti-sense primer：CGGCAAGAACTACGAGGTTT
The PCR reagent used in qualification process is as follows：
PCR reagent group 1 includes：Primer sets 1 and general PCR reagent
PCR reagent group 2 includes：Primer sets 2 and general PCR reagent
PCR reagent group 3 includes：Primer sets 3 and general PCR reagent
PCR reagent group 4 includes：Primer sets 4 and general PCR reagent
PCR reagent group 5 includes：Primer sets 5 and general PCR reagent
3. third object of the present invention, it is desirable to provide 5 in screening of the hop varieties of 10 genome sequencings
SNP site Genotyping situation (such as table 1), in case follow-up hops sample amounts and qualitative detection.
1 10 hop varieties of table are in the 5 SNP site parting situations filtered out
4. fourth object of the present invention, it is intended to hops SNP site is screened by completion, builds hops SNP site
Fingerprint database.
5. the principle of the present invention is：By carrying out full-length genome to 10 representative both at home and abroad hop varieties
Sequencing, sequencing result is contrasted to the hops draft genome issued on NCBI, obtains original SNP site.Again by original SNP positions
Point is by MIS and the filtering of minimum gene frequency, experimental reliability filtering, correlation and conspicuousness filtering, finally according to sieve
The preferable SNP site both sides sequences Design KASP specific primers of polymorphism selected, utilize the fluorescence real-time quantitatives of ABI 7500
PCR instrument enters performing PCR detection to hops sample, and the kind and purity of hops sample are judged according to the fluorescence intensity of PCR primer
Qualification result.
6. the advantage of the invention is that：Because hops complete genome group is not announced, only covered on NCBI at present
The gene sketch of 80% Genome Size.Therefore, the present invention to 10 kinds of domestic and international representative hop varieties by entering
Row high-flux sequence, contrast draft genome takes the original sites of first-hand SNP, then has been filtrated to get by multi-layer data polymorphic
The preferable SNP site of property.The mirror of kind and purity is carried out to hops sample using the fluorescence real-time quantitative PCR instrument of ABI 7500
It is fixed, it is easy to operate, result is accurate, cost is relatively low.Therefore, compared to other identification hop varieties and the detection method of purity, make
With SNP site provided by the invention and application can more directly, more accurately obtain testing result.
Brief description of the drawings
See the genotyping result of Fig. 1 Cascade samples SNP site 2.
Embodiment
Experimental method used in following embodiments is conventional method unless otherwise specified.
Material used, reagent etc., unless otherwise specified, are commercially obtained in following embodiments.
One, hops sample gene groups DNA is extracted
A collection of 48, the Cascade hop pellets sample being collected on the market is taken, is crushed respectively using liquid nitrogen grinding more
After secondary, it is added in 2ml centrifuge tubes, uses Qiagen DNeasy Plant Mini Kit (250) plant DNA extraction kit
To it by step extraction genomic DNA.
By using 1% agarose gel electrophoresis and Nanodrop 2000 to the matter of the hops genomic DNA extracted
Amount and concentration are detected, it is ensured that A260/280 is between 1.8~2.0, and A260/230 is between 1.8~2.0, and really
Hsp gene group DNA bands are completely without hangover.According to the requirement of the classifying method of the KASP specific primers of LGC companies of Britain, DNA
Dosage is 10~50ng, i.e., hops DNA concentration is diluted into 10~50ng/ μ l, standby.
Two .KASP specific PCRs react
According to the form below (such as table 2) configures PCR reaction systems, and it is glimmering that the reaction system prepared is added separately into ABI 7500 afterwards
In light real-time PCR loading wells.
Table 2KASP parting PCR reaction systems
Fluorescence real-time quantitative PCR reaction condition such as following table (such as table 3)：
The fluorescence real-time quantitative PCR reaction condition of table 3
Three, genotypic results
In the example, Cascade samples use the parting testing result such as Figure of description of SNP site 2, upper left corner round dot in figure
" GG " homozygous genotype is represented, lower right corner round dot represents " TT " homozygous genotype, and middle round dot represents " GT " heterozygous genotypes, left
Inferior horn round dot represents that " NTC " is water control sample, and the genotype is judged by detecting two kinds of fluorescence intensities in PCR primer.Figure
In each round dot be to represent the genotype of each particle Cascade hops samples, therefore the purity of this batch of Cascade sample is
38/48=79.2%, it was demonstrated that this batch of Cascade sample, which exists, to be mixed.If wanting to improve experimental result precision, sampling number can be improved
And repeat batch.
Claims (5)
1. 5 SNP sites in the hops genome that the present invention filters out, it is characterised in that can be used in following application：
Hop varieties Genotyping；
Build hops SNP site fingerprint database；
Hops sample purity Quantitative measurement；
Hops sample mixes kind Qualitative Identification；
5 SNP sites are as follows：
SNP site 1 is the Contig LD161349.1 submitted on NCBI the 9970th nucleotides, and specific place sequence is as follows：
GCCACCTACAACAATTCAAGGTGCCAGAAGGACACTTGGATCTCAGCGAACCAGGTCCTGAGGGCGCCCACATAAGA
ATATCAGCCACTCTGTCAGGAAAACAACTCCT(A/C)GTATGCAGTTGCTATTTGGAGGGCAAACACCCCAAGTTGA
ACTACAACGAGTTAATCAGATAGCTCGGGAAAACAGAGACGGACTCCCTTCTAATCCGAC；
SNP site 2 is the Contig LD205009.1 submitted on NCBI the 1776th nucleotides, and specific place sequence is as follows：
CTCGTGGAGAGTGAGACCACGCGGTTCTAAGAACACGCGCCTGTGCTCCCAACAAATCCCCAATTACTCGGAA
TTCGTGTGTCAGGAGGGTCAGGTTAGAATTTT(G/T)CCTTGGAGGGAAAGTTCCAATAGGCAAGAAAGGCAAAACC
GCCTGTGAAGCGTGTCCCCTAAGCTACCTGGTTTTGGACCCAAAATACTGGAAA；
SNP site 3 is the Contig LD169680.1 submitted on NCBI the 2676th nucleotides, and specific place sequence is such as
Under：
GTCAGGAGTTCTCAGATTACAGCTGCAAGAAGAAAGGACAGGTAAGAGAAGAATTGATCTGAAGCCGCAA
AGAAGCTAATGAAGAGCGTCTGAAG(A/G)ATTGAAAGCATAACAAGACTGGTAAGCGCGTCATCTATTGCACAAGC
ACCTCCGGAGACGACTACATCAGAGATGAGAAGAACAGCAAAACTTG；
SNP site 4 is the Contig LD167190.1 submitted on NCBI 16972 nucleotides, and specific place sequence is as follows:
TCCCGCAAGGTTCGACCCATCCTCGCGCCAGATGCGCCTGGGCGTGCACAATCCGTGACGCACTCCCACAAGT
CCATGTCCTCATCATAGA(T/C)CACGTCCTTTATGCCACCACTCTTAACTTAACCATAACACACATATATTCCAGC
ATCATCAGATGTCAATCAATTCTTACCTTGTCTTCTGAATTTCT；
SNP site 5 is the Contig LD201072.1 submitted on NCBI 2650 nucleotides, and specific place sequence is as follows：
GATATTACCAGTTCGCTGACATCTATCACAGGCCTTGACAAAAACATTAGCATCCTTGAATAATGAAGGCCAA
TAGAAACCACTTTGTAATACCTTGGC(T/C)GCCGTTCTTGATGCTCCAAAGTGCCCACCACATTGTTGAGTGTGAC
AATGGTTGAGAATTGACTGCATCTCTTCTTCAGGAACACACCTTCTGATAATCT。
2. application according to claim 1, it is characterised in that 5 SNP sites of identification hop varieties and purity are made
5 groups of primers are as follows：
(1) primer sets 1 for detecting hops genome SNP site 1 are：
Sense primer 1：CTGTCAGGAAAACAACTCCTA
Sense primer 2：CTGTCAGGAAAACAACTCCTC
Anti-sense primer：CATACGTCAACGATAAACCT
(2) primer sets 2 for detecting hops genome SNP site 2 are：
Sense primer 1：GAGGGTCAGGTTAGAATTTTG
Sense primer 2：GAGGGTCAGGTTAGAATTTTT
Anti-sense primer：GGAACCTCCCTTTCAAGGTT
(3) primer sets 3 for detecting hops genome SNP site 3 are：
Sense primer 1：CTAATGAAGAGCGTCTGAAGA
Sense primer 2：CTAATGAAGAGCGTCTGAAGG
Anti-sense primer：TAACTTTCGTATTGTTCTGA
(4) primer sets 4 for detecting hops genome SNP site 4 are：
Sense primer 1：GTCCATGTCCTCATCATAGAT
Sense primer 2：GTCCATGTCCTCATCATAGAC
Anti-sense primer：GTGCAGGAAATACGGTGGTG
(5) primer sets 5 for detecting hops genome SNP site 5 are：
Sense primer 1：CCACTTTGTAATACCTTGGCT
Sense primer 2：CCACTTTGTAATACCTTGGCC
Anti-sense primer：CGGCAAGAACTACGAGGTTT.
3. application according to claim 1, it is characterised in that the 5 hops SNP sites identification hops product filtered out
Used PCR reagent is when kind and purity：
PCR reagent group 1 includes：Primer sets 1 and general PCR reagent
PCR reagent group 2 includes：Primer sets 2 and general PCR reagent
PCR reagent group 3 includes：Primer sets 3 and general PCR reagent
PCR reagent group 4 includes：Primer sets 4 and general PCR reagent
PCR reagent group 5 includes：Primer sets 5 and general PCR reagent.
4. application according to claim 1, it is characterised in that base is carried out to hop varieties using 5 SNP site information
Because of parting, hops SNP fingerprint databases are built.
5. application according to claim 1, it is characterised in that it is qualitative fixed to be carried out using 5 SNP sites to hops sample
The step of amount detection, includes：After carrying out DNA extractions to hops sample, existed using PCR reagent group and corresponding PCR conditions
The enterprising performing PCR of real-time PCR, judges Classification Identification result by comparing fluorescence intensity afterwards.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710279239.0A CN107446991A (en) | 2017-04-25 | 2017-04-25 | A set of SNP site and its application for being applied to identification hop varieties and purity |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710279239.0A CN107446991A (en) | 2017-04-25 | 2017-04-25 | A set of SNP site and its application for being applied to identification hop varieties and purity |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN107446991A true CN107446991A (en) | 2017-12-08 |
Family
ID=60486459
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710279239.0A Pending CN107446991A (en) | 2017-04-25 | 2017-04-25 | A set of SNP site and its application for being applied to identification hop varieties and purity |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107446991A (en) |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108660247A (en) * | 2018-05-24 | 2018-10-16 | 武汉市农业科学院 | A kind of primer SmemboI-2 of the imperial No. three eggplant Purities of purple based on SNP marker and application |
| CN108676903A (en) * | 2018-05-24 | 2018-10-19 | 武汉市农业科学院 | A kind of primer SmemboI-1 of the imperial No. three eggplant Purities of purple based on SNP marker and application |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105368830A (en) * | 2015-11-19 | 2016-03-02 | 中国农业科学院棉花研究所 | Core SNP markers developed based on KASP (competitive allele specific) technology and applied to cotton hybrid identification |
-
2017
- 2017-04-25 CN CN201710279239.0A patent/CN107446991A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105368830A (en) * | 2015-11-19 | 2016-03-02 | 中国农业科学院棉花研究所 | Core SNP markers developed based on KASP (competitive allele specific) technology and applied to cotton hybrid identification |
Non-Patent Citations (2)
| Title |
|---|
| HENNING等: "Simple SNP-based minimal marker genotyping for Humulus lupulus L. identification and variety validation", 《BMC RES NOTES》 * |
| 匡猛: "基于单拷贝SNP标记的棉花杂交种纯度高通量检测技术", 《棉花学报》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108660247A (en) * | 2018-05-24 | 2018-10-16 | 武汉市农业科学院 | A kind of primer SmemboI-2 of the imperial No. three eggplant Purities of purple based on SNP marker and application |
| CN108676903A (en) * | 2018-05-24 | 2018-10-19 | 武汉市农业科学院 | A kind of primer SmemboI-1 of the imperial No. three eggplant Purities of purple based on SNP marker and application |
| CN108676903B (en) * | 2018-05-24 | 2021-06-11 | 武汉市农业科学院 | Primer SmemboI-1 for identifying purity of solanum torvum third eggplant based on SNP marker and application |
| CN108660247B (en) * | 2018-05-24 | 2021-06-11 | 武汉市农业科学院 | Primer SmemboI-2 for identifying purity of solanum torvum third eggplant based on SNP marker and application |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| García-Beneytez et al. | Application of a DNA analysis method for the cultivar identification of grape musts and experimental and commercial wines of Vitis vinifera L. using microsatellite markers | |
| CN113832243B (en) | Core SNP markers for identification of tea varieties based on KASP technology | |
| KR100956323B1 (en) | Multiple Real-Time PCR Methods for Identifying Rice Varieties | |
| CN105154544A (en) | Organism identity authentication method and biological identity authentication system based on gene detection | |
| CN107354202B (en) | Primer combination and kit for identifying flue-cured tobacco K326, application and identification method | |
| CN102321752B (en) | Fluorescence labeled detection kit for simultaneously analyzing 17 gene loci of canine genomic DNA, detection method and application thereof | |
| CN107446991A (en) | A set of SNP site and its application for being applied to identification hop varieties and purity | |
| KR101229271B1 (en) | Method of discriminating Korean native Prunus mume cultivars using SSR markers | |
| KR20120078193A (en) | Multiplex real time pcr method for discrimination of rice cultivar | |
| CN111944922B (en) | EST-SSR primer set developed based on Paeonia lactiflora transcriptome sequence and its application | |
| KR101286989B1 (en) | Multiplex real time pcr method for discrimination of rice cultivar | |
| CN113789407B (en) | A kind of SNP molecular marker combination and its application for the genotyping of oily bean | |
| KR101699518B1 (en) | Primer set for discrimination of a ginseng cultivar Gumpoong and a landrace Hwangsook and uses thereof | |
| CN112695031B (en) | Method for identifying tomato variety DNA fingerprint and SNP primer combination used by same | |
| CN109486988A (en) | The method and its kit of high throughput detection Maize Resistance To Stalk Rot Genotyping | |
| CN104593510A (en) | Primer, probe and method for detecting SNP allele variation of peanut FAD2 genes | |
| CN107446992A (en) | A set of SNP site and its application for being applied to identification sorghum variety and purity | |
| CN109852704B (en) | Composite amplification kit for simultaneously detecting 32Y chromosome loci | |
| CN104152568B (en) | High-throughput STR sequence core repeat number detection method | |
| CN107354203B (en) | Primer combination and kit for identifying cured tobacco Bina No. 1, application and detection method | |
| CN110846434A (en) | Primer and kit for identifying tea tree varieties and identification method thereof | |
| CN107354205B (en) | Primer combination and kit for identifying tobacco 100 in flue-cured tobacco, application and detection method | |
| KR102393484B1 (en) | Molecular marker based on nuclear genome sequence for discriminating genotype of Peucedanum japonicum resources and uses thereof | |
| CN110423826A (en) | A kind of C57BL/6 subbreed mouse KASP genetic detection kit and primer | |
| KR20140021696A (en) | Identifying method of diatoms in southern sea of korea, polynucleotide probe, dna chip and kit for identifying the same |
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 | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20171208 |
|
| WD01 | Invention patent application deemed withdrawn after publication |