CN110373487A - One kind InDel label relevant to capsicum pungent character and its application - Google Patents
One kind InDel label relevant to capsicum pungent character and its application Download PDFInfo
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- CN110373487A CN110373487A CN201910336087.2A CN201910336087A CN110373487A CN 110373487 A CN110373487 A CN 110373487A CN 201910336087 A CN201910336087 A CN 201910336087A CN 110373487 A CN110373487 A CN 110373487A
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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
- C12Q2600/13—Plant traits
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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 invention discloses a kind of InDel label relevant to capsicum pungent character and its applications, disclose insertion/deletion segment relevant to pepper fruit pungent character, for its nucleotide sequence as shown in SEQ ID NO.1, the fragment sequence is located at regulation capsaicine biosynthesis key myb transcription factor Cap1 upstream region of gene；It is based on this simultaneously, develops a kind of InDel molecular labeling, by detecting Cap1 upstream region of gene SEQ ID NO:1 sequence presence or absence, good parting can be carried out to Capsaicinoids content height.The present invention improves breeding efficiency for the genetic improvement of capsicum pungent, and saving breeding cost has very important directive significance.
Description
Technical field
The present invention relates to pepper breeding technical fields, more particularly, to a kind of InDel relevant to capsicum pungent character
Label and its application.
Background technique
Capsaicine and Dihydrocapsaicin, and the like to be referred to as Capsaicinoids be the distinctive one kind of capsicum plants
Secondary metabolite is the important component part of Fruit Quality of Hot Pepper, determines the pungent of fruit.Capsaicinoids food,
Medical (analgesic, anticancer, weight-reducing, blood pressure lowering, sterilization and strong stomach etc.), military (raw material for making tear bombnoun and defensive weapon) are changed
The fields such as work have extensive use.Currently, in the world to about 70,000 tons or so every year of the market demand of capsaicine, year demand it is total
Up to 17,500,000,000 yuan of value, and world's total productive capacity is annual less than 0.5 ten thousand tons, demand gap is up to 80% or more.Capsaicine class object
Matter needs are extracted from pepper fruit, and the cost for producing in extraction process 90% or more is cost of material, therefore, using capsaicine
Content is high, and the kind of disease-resistant high yield just becomes the key problem in technology of Capsaicinoids extraction.In 5 common at present pepper cultivations
In kind C.chinense Capsaicinoids content be significantly higher than other 4 kinds (C.annuum, C.baccatum,
C.frutescens,C.pubescens).However, the generally existing breeding cycle of C.chinense kind is long, percentage of fertile fruit is low, environment is suitable
Should be able to power it is weak the features such as.C.annuum kind percentage of fertile fruit is high, wide in variety, strong environmental adaptability, the wide (pepper cultivation of cultivated area
The 85% of area is this kind), but this kind of capsaicin content is low, therefore uses intraspecific hybridization strategy to this kind to improve offspring capsicum
Cellulose content ability is very limited.C.chinense and C.annuum affiliation is close, will be controlled by conventional cross-breeding method
The genetic locus of the high capsaicin content of C.chinense material of system be transferred to C.annuum material obtain high peppery, high yield, it is high-quality,
Adaptable germ plasm resource is that breeding is suitble to capsaicine processing and extracts the important prerequisite of new varieties.However traditional breeding side
That there are phenotypic evaluations is unstable, time-consuming, laborious for method, spends the disadvantages of high.
And the method for traditional detection Capsaicinoids mainly first extracts the capsaicine substance in capsicum, then uses liquid
The methods of phase chromatography, mass spectrum are measured, not only time-consuming and laborious, and at high cost, low efficiency.And isolated using objective trait
Molecular labeling, which carries out marker assisted selection, has many advantages, such as efficient, quick, stabilization, can shift to an earlier date in seedling stage to objective trait
Identification, therefore traditional conventional breeding period can be overcome long to a certain extent, land occupation is big, spends many restrictions such as more.Insertion/
Deletion polymorphism (insertion/deletion, InDel) is since the DNA sequence dna at allele site is between Different Individual
The length polymorphism variation that the insertion/deletion of nucleotide fragments has occurred and generates.In whole gene group, InDel label
Frequency of polymorphism is only second to SNP marker, is much higher than SSR marker.InDel label polymorphism can pass through polymerase chain reaction
(PCR) and the simple step such as agarose gel electrophoresis or native polyacrylamide gel electrophoresis reaches the mesh of Genotyping
's.Patent CN201811285982.8 discloses a kind of InDel label of capsicum purity of hybrid, patent
CN201810360454.8 and CN201810499212.7 discloses an InDel for distinguishing five pepper cultivation kinds and marks
Note；However it yet there are no the relevant report of the molecular labeling for distinguishing Capsaicinoids content height.
Summary of the invention
It is an object of the invention to overcome above-mentioned deficiency in the prior art, provide a kind of relevant to capsicum pungent character
InDel label.
Another object of the present invention is to provide the primers for detecting the InDel label.
A further object of the present invention is to provide a kind of methods for screening capsaicin content height character.
Above-mentioned purpose of the invention is to give realization by the following technical programs:
A kind of insertion/deletion segment relevant to pepper fruit pungent character, nucleotide sequence such as SEQ ID NO.1 institute
Show, length 66bp, is located at the gene coding region Cap1 upstream.
SEQ ID NO.1:AAACGCCTCACATCAAACATGCAATTCAAGTTCCAACATTGACAGTGCAAT TCACA
ACAACAAGAA。
The research of the invention finds that 740 self-mating systems (C.chinesne) and low capsaicin content in high capsaicin content
There is a large amount of SNP and 7 InDel for the gene coding region the Cap1 upstream sequence of CA1 self-mating system (C.annuum), wherein being most worth
It is 66bp, sequence information insertion/deletion segment as shown in SEQ NO.1, in 740 self-mating systems that length, which must be concerned with,
In the presence of and being missing from CA1 self-mating system, and from the C.chinense genes of 47 representational high capsaicin contents
Profile material and 73 representational C.annuum Genotypes analyze Cap1 upstream of coding region sequence, find the 66bp
Segment all exists in all C.chinense Genotypes, and then lacks in C.annuum Genotype, shows SEQ ID
Insertion/deletion segment shown in NO.1 is related to the pungent character of capsicum, can be used for the screening of capsaicin content height character, uses
In the peppery degree of fruit of prediction capsicum plant body or the peppery degree of fruit of its filial generation.Show to can also be used shown in SEQ ID NO.1 simultaneously
Insertion/deletion segment distinguish C.annuum Genotype and C.chinense Genotype.
Therefore, the present invention be also claimed the insertion/deletion segment in screening capsaicin content height character or
Application in the kit of preparation screening capsaicin content height character；And the insertion/deletion segment is distinguishing capsicum
Application in C.annuum Genotype and C.chinense Genotype.
The present invention also provides a kind of for screening the InDel molecular labeling of capsaicin content height character, including upstream is drawn
Object and downstream primer, sequence is successively as shown in NO.3~4 SEQ ID:
InDel-F:5 '-CGCTTCCTACTGGTAGAATCAA-3 ' (SEQ ID NO.3)；
InDel-R:5 '-TTCTGCTGTGGATAATGATTAG-3 ' (SEQ ID NO.4).
The present invention is also claimed the InDel molecular labeling and sieves in screening capsaicin content height character or in preparation
Select the application in the kit of capsaicin content height character.
A kind of method of the Capsaicinoids content height of screening/prediction capsicum or its filial generation, includes the following steps:
S1. capsicum plant body genomic DNA to be measured is extracted；
S2. Cap1 upstream region of gene whether there is insertion/deletion shown in SEQ ID NO:1 in genome described in detecting step S1
Segment；
S3. result judgement: if there are insertion/deletion segments shown in SEQ ID NO.1 in genome, then it represents that be measured peppery
Capsaicinoids content is high in green pepper, and peppery degree is high；If not existing, then it represents that Capsaicinoids content is lower, and peppery degree is low.It can establish
The method of seedling stage molecular labeling assisting sifting capsaicin content height character.
Or: if there are insertion/deletion segments shown in SEQ ID NO.1 in genome, then it represents that capsicum to be measured is
C.chinense Genotype；If it does not exist, then it represents that capsicum to be measured is C.annuum Genotype.
The capsicum plant body be include but is not limited to the root of pepper plant, stem, leaf, flower, pericarp, seed or other can
Extract position, tissue or the derived tissues of capsicum complete genome group.
Preferably, it is detected as described in step S2 using genomic DNA described in step S1 as template, with above-mentioned InDel molecule
Label carries out pcr amplification reaction；Capsicum to be measured or the capsaicine that its filial generation contains high level are indicated if obtaining 434bp segment
The peppery degree of substance is high；Indicate that Capsaicinoids content to be measured is lower if there is 368bp band, peppery degree is low.
Preferably, the reaction system of the PCR are as follows: 10 μ L Taq enzyme PCR Mix, each 1 μ L of upstream and downstream primer, DNA profiling
50ng adds ddH2O to 20mL.
Preferably, the response procedures of the PCR are as follows: 95 DEG C of initial denaturation 10min；95 DEG C of denaturation 10s, 55 DEG C of annealing 10s, 72
DEG C extend 30s, repeat 35 circulation；72 DEG C of extension 10min.
The present invention also provides a kind of for screening/predicting the kit of the Capsaicinoids of capsicum or its filial generation, includes
For detecting the primer pair of insertion/deletion segment described in SEQ ID NO.1.
Preferably, the primer pair includes upstream primer and downstream primer, and sequence is successively such as SEQ ID NO.3~4 institute
Show.
InDel molecular labeling shown in insertion/deletion segment shown in SEQ ID NO:1 of the present invention or NO.3~4 SEQ ID exists
Application in capsicum assistant breeding is also in the scope of the present invention.
Compared with prior art, the invention has the following advantages:
The present invention provides a kind of insertion/deletion segments relevant to pepper fruit pungent character, and nucleotide sequence is such as
Shown in SEQ ID NO.1, the fragment sequence is located on regulation capsaicine biosynthesis key myb transcription factor Cap1 gene
Trip；Based on this, a kind of InDel molecular labeling is developed, sequence successively as shown in NO.3~4 SEQ ID, passes through detection Cap1
Upstream region of gene SEQ ID NO:1 sequence presence or absence can carry out good parting to Capsaicinoids contents height, can be
Plant strain growth early stage can predict the height of capsaicin content；It can be rejected early in breeding process as a result, and not conform to breeding objective
Plant, avoid unnecessary artificial and fund cost waste.The present invention improves breeding for the genetic improvement of capsicum pungent
Efficiency, saving breeding cost has very important directive significance.
Detailed description of the invention
Fig. 1 is the identification of Capsaicinoids content main effect QTL site Cap1；Figure a represents 740 and CA1 filial generation F2
The distribution of group's capsaicin content；Figure b represents F2Group's Dihydrocapsaicin content distribution；Figure c represents F2The total capsaicin content of group point
Cloth；Figure d represents capsaicin content QTL positioning；Figure e represents Dihydrocapsaicin content QTL positioning；Figure f represents total capsaicin content
QTL positioning.
Fig. 2 is Cap1 finely positioning.
Fig. 3 is the analysis of the gene coding region Cap1 upstream sequence and marker development.
Fig. 4 is to be analyzed using label material Capsaicinoids content.
Fig. 5 analyzes filial generation capsaicin content using label；Scheming a indicates 2013 Guiness World Records report
Extremely capsicum " Caro Lehner Death capsicum " (Carolina Reapers) photo；Scheming b indicates Cap1 upstream region of gene 66bp sequence
It is listed in " Caro Lehner Death capsicum " (Carolina Reapers) detection；Scheming c indicates to use InDel labeled analysis Carolina
Reapers and CA filial generation F2Group's loci gene type；Scheming d indicates filial generation F2Group's Cap1 upstream region of gene contains
The corresponding capsaicine of Carolina Reapers or CA sequence single plant (on) and Dihydrocapsaicin (under) content.
Specific embodiment
Principles and features of the present invention are described below in conjunction with example, the given examples are served only to explain the present invention, and
It is non-to be used to limit the scope of the invention.According to description below and these embodiments, those skilled in the art can determine this hair
Bright essential characteristic, and without departing from the spirit and scope of the invention, the present invention can be made various changes and
Modification, so that it is applicable in various uses and condition.Experimental method in following embodiments is unless otherwise specified conventional side
Method, such as sambrook molecular cloning experiment handbook (Sambrook J&Russell DW, Molecular cloning:a
Laboratory manual, 2001), or the condition of the specification suggestion according to biochemical reagents manufacturer.In following embodiments
Material, reagent used etc. commercially obtain unless otherwise specified.
The acquisition of 1 InDel of embodiment label
Make maternal and low capsaicin content CA1 using with 740 self-mating systems (C.chinesne) of high capsaicin content
Self-mating system (C.annuum) makees male parent, carries out hybridization and obtains F1Generation, later by F1Generation selfing obtains F2Group.By 150 F2Group
Greenhouse is colonized in later using hole plate seedling growth and carries out Routine Management.Group's single plant SNP is measured using SLAF technology, develops one
It is a to contain 9038 highdensity genetic maps of label.The extraction and measurement of the capsaicine and Dihydrocapsaicin of group are using efficient
Liquid chromatography.The main effect QTL site Cap1 of Capsaicinoids content is navigated to 7 using composite interval mapping method (CIM)
Number chromosome, the QTL can explain 37~42% phenotypic variations (Fig. 1).According to the site Cap1 corresponding position on chromosome
Corresponding InDel label is developed, further using CA1 self-mating system as recurrent parent and F1Cap1 is limited to 510kb by generation hybridization
Range, which contains 6 genes and all encodes myb transcription factor, and wherein Cap1 is by we demonstrate that directly take part in peppery
The biosynthesis (Fig. 2) of green pepper element, the nucleotide sequence of Cap1 gene is as shown in SEQ ID NO.2.
By further cloning the gene coding region the Cap1 upstream sequence of capsicum 740 and CA1 self-mating system, hair is between the two
There are a large amount of SNP and 7 InDel, wherein most it is worth noting that 66bp sequence as shown in SEQ NO:1,740 from
It hands in system (Fig. 3) for existing, and being missing from CA1 self-mating system, and from 47 representational high capsaicin contents
C.chinense Genotype and 73 representational C.annuum Genotypes analyze Cap1 upstream of coding region sequence
Column, discovery 66bp segment all exists in all C.chinense Genotypes, and then lacks in C.annuum Genotype.
A pair of of InDel label SEQ ID NO.3 and SEQ ID NO.4 is developed based on this.The molecular labeling can be educated in capsicum annuum
It is applied in kind.Method particularly includes: capsicum genomic DNA is used, is carried out using SEQ ID NO.3 and SEQ ID NO.4 primer
PCR amplification, amplified production electrophoresis on 2% agarose gel prove to contain high Capsaicinoids if obtaining 434bp segment
Content sequence, the Capsaicinoids of material (filial generation) containing high level.
Embodiment 2
Capsaicinoids (capsaicine and Dihydrocapsaicin) content is using efficiently measurement.The sample of acquisition is placed on 40 DEG C
48h is dried in baking oven, is ground while hot with mortar.Ground sample 0.1g is weighed, is placed in the centrifuge tube of 15mL, writes respectively
Label.The extracting solution (methanol: tetrahydrofuran (1:1)) of 10mL is added in each centrifuge tube, covers lid in supersonic wave cleaning machine
Then ultrasonic 30min stands overnight sample room temperature.1mL Capsaicinoids extracting solution, warp are drawn with disposable syringe
0.22 μm of membrane filtration is packed into 1mL chromatogram bottle and carries out chromatography.Draw 10 μ L standard specimens or sample to be tested high-efficient liquid phase color
Spectrometer WatersAlliance 2489 by XSelect HSS C-18SB column (4.6 × 250mm, 5 μm) chromatographic column into
Row separation, the sample after separation carry out absorption blob detection with ultraviolet detection system 2489UV/Visible at 280nm.Chromatostrip
Part are as follows: 80% methanol flow rate 1mL/min, 30 DEG C of post case temperature, each sample analysis time 10min.Capsaicine and dihydro capsicum
Cellulose content is calculated according to the standard curve that standard items make, and total Capsaicinoids content is according to (capsaicine+dihydro capsicum
Element)/0.9 calculated (Bennett and Kirby, 1968).
11 representational C.chinense Genotypes are extracted using CTAB method and 10 representational
The DNA of C.annuun Genotype.Using the genomic DNA of extraction as template, with InDel primer SEQ ID NO.3 and SEQ
ID NO.4 carries out PCR amplification.Amplification system is 20 μ L, and included component is as follows: 10 μ L Taq enzyme PCR Mix, upstream and downstream primer
Each 1 μ L, DNA profiling 50ng, later plus distilled water H2O polishing is to 20mL.Using Bio-Rad T100TMPCR carries out PCR amplification,
Its amplification program is as follows: 95 DEG C of initial denaturation 10min, then 95 DEG C denaturation 10s, 55 DEG C annealing 10s, 72 DEG C extension 30s program
Carry out 35 circulations.5 μ L reaction PCR product is taken to carry out electrophoresis, electrophoresis 40min, is contaminated with EB later on 2% agarose
Color 15min, and taken pictures using Bio-Rad Gel Doc XR+Gel Doc XR gel imaging system.According to the item of reading
Band and Marker are compared, and have been found that then the material Capsaicinoids content is higher there are 434bp band, if there is
Then the Capsaicinoids content is lower (Fig. 4) for 368bp band, consistent with the testing result of high performance liquid chromatography.
Embodiment 3
Made using most peppery kind ' the Carolina Reaper ' (C.chinesne) of 2013 Guiness World Records report female
This and micro- peppery CA1 self-mating system (C.annuum) make male parent, carry out hybridization and obtain F1Generation, later by F1Generation selfing obtains F2Group,
It is colonized and carries out conventional cultivation management.F2Group DNA extraction, Capsaicinoids assay, PCR amplification and band inspection
It surveys referring to described in case study on implementation 2.According to the band of reading, have been found that there are 434bp band then offspring's materials in filial generation
Expect Capsaicinoids content it is higher, if there is 368bp band then the progeny material Capsaicinoids content it is lower (figure
5)。
It is had confirmed by above-mentioned experiment, capsicum is detected by InDel primer SEQ ID NO.3 and SEQ ID NO.4
Identify sequence shown in SEQ ID NO:1 in plant genome, can effectively predict the capsaicine class object of different genotype capsicum
Matter content., can be after the completion of hybridization using this feature, plant strain growth early stage can predict the height of capsaicin content.As a result,
The plant for not conforming to breeding objective can be rejected early in breeding process, avoid unnecessary artificial and fund cost waste.
The foregoing is merely presently preferred embodiments of the present invention, is not intended to limit the invention, it is all in spirit of the invention and
Within principle, any modification, equivalent replacement, improvement and so on be should all be included in the protection scope of the present invention.
Sequence table
<110>Agricultural University Of South China
<120>a kind of InDel label relevant to capsicum pungent character and its application
<141> 2019-04-24
<160> 4
<170> SIPOSequenceListing 1.0
<210> 1
<211> 66
<212> DNA
<213>capsicum (Capsicum annuum L.)
<400> 1
aaacgcctca catcaaacat gcaattcaag ttccaacatt gacagtgcaa ttcacaacaa 60
caagaa 66
<210> 2
<211> 750
<212> DNA
<213>capsicum (Capsicum annuum L.)
<400> 2
atggtgagaa caccttgcta cgacgaaaat ggaatgaaga aggggacatg gactcctgaa 60
gaagatagga agttaacagc atatattgca aaatatggct catggaactg gcgccaactt 120
cccaagtatg ctggactagc aaggtgtgga aagagctgca gacttcgatg gatgagtcac 180
ttacggccaa atgttaaaag agggaattat accaaagaag aagatgaaat catcttgaac 240
ctccatgctc aacttggaaa taggtggtcg gcgattgctg ctcacttgcc aggaagatca 300
gacaatgaga taaagaatca ttggcacaca aaacttaaga agcgcggtac taattatgcg 360
acaaactcaa gtgatgaatc aagcaagaaa tgtaagaata atactaagaa gaggtatact 420
gaaagtaata ccaataaaaa tacaagtcat aataatatgc aggaaaatat agtactggaa 480
agtttagaat ggtcaccaaa ggaatcatca agtgaagaac tctcctctta cagtaccact 540
aattatcaac agcaacataa agtgtttcaa gaggaaataa ctagtggaag cttttggaca 600
gaaccatttg tagtagaaag tttcaatact actagaactg attttctagc tccttcaatt 660
gattactgtg gacttgtgtg tccaccttca ccatatatag gtcatgaatt tctttcctcc 720
tttgactttg atcattataa ttattggtaa 750
<210> 3
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<212> DNA
<213>capsicum (Capsicum annuum L.)
<400> 3
cgcttcctac tggtagaatc aa 22
<210> 4
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<212> DNA
<213>capsicum (Capsicum annuum L.)
<400> 4
ttctgctgtg gataatgatt ag 22
Claims (10)
1. a kind of insertion/deletion segment relevant to pepper fruit pungent character, which is characterized in that its nucleotide sequence such as SEQ
Shown in ID NO.1.
2. insertion/deletion segment described in claim 1 screens capsaicine in screening capsaicin content height character or in preparation
Application in the kit of content height character.
3. a kind of for screening the InDel molecular labeling of capsaicin content height character, which is characterized in that including upstream primer and
Downstream primer, sequence is successively as shown in NO.3~4 SEQ ID.
4. the method for the Capsaicinoids content height of a kind of screening/prediction capsicum or its filial generation, which is characterized in that including such as
Lower step:
S1. the genomic DNA of capsicum to be measured is extracted；
S2. with the presence or absence of insertion/deletion segment shown in SEQ ID NO.1 in genome described in detecting step S1；
S3. result judgement: if there are insertion/deletion segments shown in SEQ ID NO.1 in genome, then it represents that in capsicum to be measured
Capsaicinoids content is high；If not existing, then it represents that Capsaicinoids content is lower.
5. according to the method described in claim 4, it is characterized in that, being detected as described in step S2 with genome described in step S1
DNA is template, carries out pcr amplification reaction with InDel molecular labeling as claimed in claim 3；It is indicated if obtaining 434bp segment
Capsicum to be measured or its filial generation contain the Capsaicinoids of high level, and capsaicine class to be measured is indicated if there is 368bp band
Content of material is lower.
6. according to the method described in claim 5, it is characterized in that, the reaction system of the PCR are as follows: 10 μ L Taq enzyme PCR
Mix, upstream and downstream primer each 1 μ L, DNA profiling 50ng add ddH2O to 20mL.
7. according to the method described in claim 5, it is characterized in that, the response procedures of the PCR are as follows: 95 DEG C of initial denaturation 10min；
95 DEG C of denaturation 10s, 55 DEG C of annealing 10s, 72 DEG C of extension 30s repeat 35 circulations；72 DEG C of extension 10min.
8. a kind of for screening/predicting the kit of the Capsaicinoids of capsicum or its filial generation, which is characterized in that comprising being used for
Detect the primer pair of insertion/deletion segment described in claim 1.
9. kit according to claim 8, which is characterized in that the primer pair includes upstream primer and downstream primer,
Its sequence is successively as shown in NO.3~4 SEQ ID.
10. InDel molecular labeling described in insertion/deletion segment or claim 3 described in claim 1 is in capsicum assistant breeding
Application.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN113151568A (en) * | 2021-05-20 | 2021-07-23 | 中国农业科学院蔬菜花卉研究所 | SNP (Single nucleotide polymorphism) site with closely linked capsaicin content, CAPS (cleaved amplified polymorphic sequence) molecular marker of SNP site and application of CAPS molecular marker |
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| CN107151709A (en) * | 2017-07-13 | 2017-09-12 | 中国农业科学院蔬菜花卉研究所 | The SNP marker related to capsicum pungent and its application |
| CN108384875A (en) * | 2018-03-23 | 2018-08-10 | 湖南省蔬菜研究所 | A kind of and the relevant insertion/deletion site of capsicum mellow fruit color gene, molecular labeling, molecular labeling primer and application |
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