CN116287297A - Litopenaeus vannamei whole genome SNP chip and application thereof - Google Patents
Litopenaeus vannamei whole genome SNP chip and application thereofInfo
- Publication number
- CN116287297A CN116287297A CN202310120282.8A CN202310120282A CN116287297A CN 116287297 A CN116287297 A CN 116287297A CN 202310120282 A CN202310120282 A CN 202310120282A CN 116287297 A CN116287297 A CN 116287297A
- Authority
- CN
- China
- Prior art keywords
- snp
- chip
- genome
- litopenaeus vannamei
- whole genome
- 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
- 241000238553 Litopenaeus vannamei Species 0.000 title claims abstract description 49
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 8
- 238000009826 distribution Methods 0.000 claims abstract description 6
- 238000004458 analytical method Methods 0.000 claims description 13
- 230000002068 genetic effect Effects 0.000 claims description 12
- 241000238557 Decapoda Species 0.000 claims description 10
- 238000005516 engineering process Methods 0.000 claims description 10
- 238000012216 screening Methods 0.000 claims description 7
- 238000000034 method Methods 0.000 claims description 6
- 239000000523 sample Substances 0.000 claims description 6
- 238000002360 preparation method Methods 0.000 claims description 4
- 238000009394 selective breeding Methods 0.000 claims description 4
- 238000009827 uniform distribution Methods 0.000 claims description 4
- 241000318927 Shrimp white spot syndrome virus Species 0.000 claims description 2
- 230000004083 survival effect Effects 0.000 claims description 2
- 230000007935 neutral effect Effects 0.000 claims 1
- 238000009395 breeding Methods 0.000 abstract description 13
- 230000001488 breeding effect Effects 0.000 abstract description 13
- 238000012098 association analyses Methods 0.000 abstract description 3
- 238000013461 design Methods 0.000 abstract description 3
- 238000011156 evaluation Methods 0.000 abstract description 3
- 238000005457 optimization Methods 0.000 abstract 1
- 238000001914 filtration Methods 0.000 description 8
- 108700028369 Alleles Proteins 0.000 description 7
- 239000003550 marker Substances 0.000 description 6
- 238000003205 genotyping method Methods 0.000 description 5
- 238000012163 sequencing technique Methods 0.000 description 5
- 241001465754 Metazoa Species 0.000 description 4
- 210000000349 chromosome Anatomy 0.000 description 4
- 239000002773 nucleotide Substances 0.000 description 4
- 125000003729 nucleotide group Chemical group 0.000 description 4
- 238000011160 research Methods 0.000 description 4
- 241000282414 Homo sapiens Species 0.000 description 3
- 241000238550 Penaeidae Species 0.000 description 3
- 230000006978 adaptation Effects 0.000 description 3
- 238000004891 communication Methods 0.000 description 3
- 238000012268 genome sequencing Methods 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000007400 DNA extraction Methods 0.000 description 2
- 241000726221 Gemma Species 0.000 description 2
- 238000009360 aquaculture Methods 0.000 description 2
- 244000144974 aquaculture Species 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 238000002373 gas-phase electrophoretic mobility molecular analysis Methods 0.000 description 2
- 238000012252 genetic analysis Methods 0.000 description 2
- 238000012214 genetic breeding Methods 0.000 description 2
- 238000012165 high-throughput sequencing Methods 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 210000003205 muscle Anatomy 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 101150084750 1 gene Proteins 0.000 description 1
- 229920000936 Agarose Polymers 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 241000252233 Cyprinus carpio Species 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 241001596950 Larimichthys crocea Species 0.000 description 1
- 240000007594 Oryza sativa Species 0.000 description 1
- 235000007164 Oryza sativa Nutrition 0.000 description 1
- 241000237502 Ostreidae Species 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 241000277263 Salmo Species 0.000 description 1
- 208000005652 acute fatty liver of pregnancy Diseases 0.000 description 1
- 238000000246 agarose gel electrophoresis Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 239000013078 crystal Substances 0.000 description 1
- 238000007405 data analysis Methods 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 230000004907 flux Effects 0.000 description 1
- 210000002683 foot Anatomy 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 230000008676 import Effects 0.000 description 1
- 230000010365 information processing Effects 0.000 description 1
- 238000007689 inspection Methods 0.000 description 1
- 101150044508 key gene Proteins 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 238000013178 mathematical model Methods 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 235000020636 oyster Nutrition 0.000 description 1
- 230000002093 peripheral effect Effects 0.000 description 1
- 238000002205 phenol-chloroform extraction Methods 0.000 description 1
- 244000144977 poultry Species 0.000 description 1
- 230000001737 promoting effect Effects 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 238000013441 quality evaluation Methods 0.000 description 1
- 235000009566 rice Nutrition 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 238000010008 shearing Methods 0.000 description 1
- 238000002791 soaking Methods 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 230000009182 swimming Effects 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
- 238000012070 whole genome sequencing analysis Methods 0.000 description 1
Images
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
-
- 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
- C12Q1/6813—Hybridisation assays
- C12Q1/6834—Enzymatic or biochemical coupling of nucleic acids to a solid phase
- C12Q1/6837—Enzymatic or biochemical coupling of nucleic acids to a solid phase using probe arrays or probe chips
-
- C—CHEMISTRY; METALLURGY
- C40—COMBINATORIAL TECHNOLOGY
- C40B—COMBINATORIAL CHEMISTRY; LIBRARIES, e.g. CHEMICAL LIBRARIES
- C40B40/00—Libraries per se, e.g. arrays, mixtures
- C40B40/04—Libraries containing only organic compounds
- C40B40/06—Libraries containing nucleotides or polynucleotides, or derivatives thereof
-
- 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
- C12Q2600/124—Animal traits, i.e. production traits, including athletic performance or the like
-
- 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
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A40/00—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production
- Y02A40/80—Adaptation technologies in agriculture, forestry, livestock or agroalimentary production in fisheries management
- Y02A40/81—Aquaculture, e.g. of fish
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Analytical Chemistry (AREA)
- Molecular Biology (AREA)
- Biochemistry (AREA)
- Immunology (AREA)
- Microbiology (AREA)
- Physics & Mathematics (AREA)
- Biotechnology (AREA)
- Biophysics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention belongs to the field of marine organism breeding, and particularly relates to a full genome SNP chip of litopenaeus vannamei and application thereof. The chip contains 594875 SNP sites, and the information of the SNP is shown in Table 1. The SNP loci of the chip are all high-quality SNP loci subjected to optimization design, have the characteristics of uniform genome distribution, high typing success rate, good representativeness and the like, can be used for positioning genes related to economic traits of the litopenaeus vannamei, whole genome association analysis, germplasm evaluation and the like, and has wide application prospects in molecular breeding of the litopenaeus vannamei.
Description
Technical Field
The invention belongs to the field of marine organism breeding, and particularly relates to a full genome SNP chip of litopenaeus vannamei and application thereof.
Background
SNP (Single nucleotide polymorphism), a single nucleotide polymorphism, is the most widely distributed marker in the genome, and compared with AFLP, RAPD of the first generation and SSR markers of the second generation, SNP markers have the characteristics of large quantity, wide range, dominant marker, direct correlation with characters, convenience in high-throughput typing and the like. SNP markers are widely applied to population genetic diversity analysis, germplasm genetic relationship identification, genetic map construction, QTL positioning, whole genome association analysis and the like. Meanwhile, based on SNP markers of whole genome, meuwissen et al proposed a whole genome selective breeding technique in 2001 (Meuwissen T H E, hayes B J, goddard M E.prediction of total genetic value using genome-wide dense marker maps. Genetics,2001,157 (4): 1819-1829.), calculated the effect of each chromosome fragment by using SNP covering the whole genome, calculated the genome breeding value of the individual by mathematical model, and then selected and remained the individual. Compared with the traditional BLUP technology, the whole genome breeding greatly improves the breeding accuracy. SNP markers therefore play an important role in genetic and breeding studies.
The key to SNP marker application is to achieve its high throughput discovery and typing. At present, the main mode of SNP (single nucleotide polymorphism) discovery is high-throughput genome resequencing or transcriptome sequencing of different samples, so far, a large number of SNP markers in livestock, poultry, crops and aquatic animals have been discovered by utilizing the technology, and a foundation is laid for analyzing species evolution and breeding paths and analyzing mechanisms of character formation. In the aspect of high-throughput SNP typing, high-throughput sequencing and gene chip are two main technical means. High throughput sequencing is divided into whole genome resequencing and simplified genome sequencing, and whole genome resequencing can obtain all SNP markers of genome, is an important technology for carrying out fine analysis and positioning of characters, but for species with larger genome and high complexity, the cost of whole genome sequencing is high, the analysis period is long, and the application in breeding is less. At present, the technology based on simplified genome sequencing such as RAD (random access memory), 2b-RAD, GBS, SLAF, targeted sequencing and the like has the advantages of low cost, high typing efficiency and the like, and is widely applied to genetic research, but the simplified genome sequencing also has short plates such as low typing mark overlapping rate, long typing period and the like among different individuals due to the complicated database building step.
The gene chip is characterized in that nucleotide probes around a target SNP are synthesized on a crystal or solid substrate, genotype information of each SNP locus is obtained through DNA hybridization, scanning and information processing, and various high-density SNP chips such as human beings, multiple-mode organisms, rice, cattle, sheep and the like are developed in the mainstream application, and various high-density SNP chips such as Atlantic salmon, carp, large yellow croaker, oyster and the like are designed in aquatic animals at present, but effective high-density SNP chips are not available in prawns.
The litopenaeus vannamei is an aquaculture economic animal which is important in China and the world, the annual output of the litopenaeus vannamei is about 500 ten thousand tons worldwide, the litopenaeus vannamei is the aquaculture economic animal with the highest output value of single variety worldwide, the Chinese is the country with the highest output of the litopenaeus vannamei, the annual output reaches 160 ten thousand tons, and the large-scale culture of the litopenaeus vannamei provides rich high-quality protein for human beings. Based on the important industrial status of Litopenaeus vannamei, genetic breeding research of Litopenaeus vannamei is carried out in the United states, thailand, australia and China. However, the current basic research of genetics of Litopenaeus vannamei is weak, the whole genome sequence is not decoded until 2019 (Zhang X, yuan J, sun Y, et al Penaeid shrimp genome provides insights into benthic adaptation and frequent Multi. Nature Communications,2019,10 (1): 356), genetic analysis of economic traits of Litopenaeus vannamei and development of modern breeding are slow. The establishment of the high-throughput SNP typing technology is a key for promoting economic character analysis and whole genome breeding technology development, and the SNP covering the whole genome can be utilized to accelerate the fine positioning of the economic character, and meanwhile, the high-throughput typing of the high-density SNP marker also provides technical support for the application of the whole genome breeding technology.
Disclosure of Invention
The invention aims to provide a litopenaeus vannamei whole genome SNP chip and application thereof.
In order to achieve the above purpose, the invention adopts the following technical scheme:
a full genome SNP chip of litopenaeus vannamei contains 594875 SNP loci, and the information of the SNP is shown in table 1.
The preparation method of the Litopenaeus vannamei whole genome SNP chip comprises the following steps:
(1) Utilizing genome-wide reference map and genome-wide variation data (Zhang X, yuan J, sun Y, et al Penaeid shrimp genome provides insights into benthic adaptation and frequent Multi Nature Communications,2019,10 (1): 356) of Litopenaeus vannamei, screening high-quality SNP loci according to MAF >0.05, and constructing an SNP database of Litopenaeus vannamei;
(2) Analyzing the sequence around the SNP locus, deleting the SNP locus which cannot be designed by the probe, and deleting loci containing other SNPs within the range of 35bp around the SNP locus;
(3) On the genome, taking 2kb as a sliding window, randomly selecting 1 SNP locus in each window to form SNP data of genome even distribution;
(4) Screening SNP loci positioned in an exon region according to the annotation information of the SNP, adding the SNP loci into the obtained genome uniform distribution SNP data, and selecting 1 SNP locus in each window according to 500bp serving as a sliding window, so that the genome exon region; meanwhile, adding the SNP locus of the splice into the SNP data set;
(5) Adding the sites related to the characters into the SNP data set, preparing the SNP data set by using a chip technology, and finally obtaining the 600K SNP typing chip of the Litopenaeus vannamei genome, wherein the SNP typing chip contains 594875 SNP sites in total.
And (3) the character growth, WSSV resistance, sex resistance, AHPND resistance and survival rate in the character related sites in the step (5).
Application of Litopenaeus vannamei whole genome SNP chip in analysis of germplasm genetic relationship and germplasm identification
The application of the Litopenaeus vannamei whole genome SNP chip in identification of associated genes of prawn characters.
The application of the Litopenaeus vannamei whole genome SNP chip in the whole genome selective breeding of the prawns.
Further, the breeding application method of the litopenaeus vannamei whole genome SNP chip specifically comprises the following steps:
(1) Extracting prawn DNA: taking tissues such as swimming feet, muscles, blood or tentacles of an analysis sample, and extracting prawn DNA by using a tenna genome DNA extraction kit.
(2) Quality inspection and standardization of DNA: the extracted DNA was assayed for DNA concentration using Nanodrop and DNA integrity was checked by 1% agarose electrophoresis, with concentrations greater than 50 and the DNA with intact main band diluted to 50 ng/. Mu.l for use.
(3) Chip typing of "middle Ke core No. 1": the extracted DNA is subjected to genotyping by using a designed SNP chip of Litopenaeus vannamei 'Zhongke core No. 1', the genotyping step is operated according to the standard flow of Affymetrix gene chip detection, and genotype data of 600K SNP loci of 96 samples are obtained through one-time detection.
(4) Data analysis: converting the obtained SNP file into a plink format, filtering SNP loci by using plink software according to the typing success rate of more than 95 percent and the minimum allele frequency of more than 0.05, and filtering individuals according to the locus typing success rate of more than 95 percent of a single individual as a standard to obtain high-quality SNP data. Genetic analysis of population PCA, GWAS, GS and the like was then performed using PLINK, GCTA, GEMMA, HIBLUP, BGLR and the like.
The whole genome reference genome information of the litopenaeus vannamei is http:// www.shrimpbase.net/vannamei.html or https:// www.ncbi.nlm.nih.gov/asembly/GCF_ 003789085.1/.
The invention has the advantages that:
(1) The high-density SNP chip of the litopenaeus vannamei provided by the invention is a chip optimally designed by genome, the genome of SNP loci is uniformly distributed, and the chromosome SNP density is high.
(2) The litopenaeus vannamei whole-gene SNP chip provided by the invention has the characteristics of high typing flux and high typing accuracy, and can realize the genotyping of 60 ten thousand SNP loci of 96 samples at one time.
Drawings
FIG. 1 is a SNP marker distribution diagram of a Litopenaeus vannamei "middle core No. 1" chip provided by the embodiment of the invention.
FIG. 2 is a diagram showing the frequency distribution of high-quality SNP minimal alleles screened by using the chip according to the invention according to the embodiment of the invention.
Fig. 3 is a graph of the analysis result of genetic relationship (PCA) of different populations using the "mesocore No. 1" of litopenaeus vannamei according to the embodiment of the present invention.
Detailed Description
The following examples are illustrative of the present invention and are not intended to limit the scope of the invention. Modifications and substitutions to methods, procedures, or conditions of the present invention without departing from the spirit and nature of the invention are intended to be within the scope of the present invention. The technical means used in the examples are conventional means well known to those skilled in the art unless otherwise indicated.
The gene chip is suitable for SNP typing of the whole genome of the litopenaeus vannamei, can realize the typing of 60 ten thousand SNP loci at the same time, and provides an important tool for economic character key gene positioning and whole genome selective breeding research of the litopenaeus vannamei.
Example 1: preparation of SNP chip of core No. 1 in Litopenaeus vannamei
1. SNP data Source
Genome re-sequencing was performed for 38 individuals of different sources including domestic and foreign germplasm, sequencing depth was 20× for each individual, obtaining high quality reads after filtering sequencing data, aligning to published litopenaeus vannamei reference genome (Zhang X, yuan J, sun Y, et al Penaeid shrimp genome provides insights into benthic adaptation and frequent mole Communications,2019,10 (1): 356) using BWA, and SNP typing using GATK software to obtain 17223769 SNP sites, which were further filtered by minimum allele frequency (> 0.05) and site typing success rate (> 0.9) to obtain 9298386 high quality SNP sites.
2. Screening of SNP loci
The type of SNP is filtered first, and the filtering criteria are as follows: (1) only the SNP sites of the alleles remain; (2) Extracting 35bp sequences on the upstream and downstream of the two-allele SNP loci, performing blast comparison on the sequences and a reference genome, and only preserving the sequences uniquely matched with the genome to ensure that the SNP loci are unique in the genome. 6372569 SNPs are obtained after the filtration, and the SNP loci and the peripheral sequences thereof are sent to a Thermofisher company for SNP locus quality evaluation so as to carry out further screening.
3. Site selection for SNP chip customization
After SNP locus evaluation, 3806943 SNP markers can be used for chip design, and SNP loci for chip design are further screened on the basis of the above, and screening is carried out according to the following standard:
(1) Deleting the loci containing other SNPs within 35bp range around the SNP locus; (2) On the genome, taking 2kb as a sliding window, randomly selecting 1 SNP site in each window, and skipping if not, so as to form SNP data with genome uniformly distributed; (3) According to the annotation information of the SNP, SNP loci in the exon region are screened out, the SNP loci annotated as selected shearing loci are added into the obtained genome uniform distribution SNP data, 1 SNP locus is selected in each window according to 500bp as a sliding window, if not, the SNP loci are skipped, and therefore the density of the SNP loci is higher in the genome exon region.
To increase the number of functional SNP sites in the genome, 2000 growth, anti-WSSV, sex-related sites screened earlier were added to the above SNP dataset to obtain 649903 SNP datasets.
4. Preparation of SNP chip
The genome SNP typing chip of the litopenaeus vannamei is prepared by using the Affymetrix chip technology of Thermofiser company, 594936 SNP probes are finally synthesized, because part of the probes are the same SNP locus, 594875 SNP loci are contained in the core No. 1 of the litopenaeus vannamei, and the distribution of the SNP on the chromosome is shown in figure 1.
Example 2: genotyping and application of Litopenaeus vannamei midkinensis core No. 1
1. Sampling and DNA extraction of different germplasm materials of prawns
Collecting 3 different sources of prawn variety materials in China, wherein A is a domestic population, B is an import population, C is an Ecuador population, and 32 individuals (each of which is known to be commercially available) are sampled from each source. Soaking muscle in 95% alcohol, extracting DNA of each individual by phenol chloroform method, detecting the integrity of the extracted DNA by agarose gel electrophoresis, detecting the concentration of the DNA by Nanodrop, and uniformly diluting to 50 ng/. Mu.l for standby.
Chip typing of "Zhongke core No. 1
The DNA of different groups is subjected to genotyping by using a 'Zhongke core No. 1', hybridized, washed and scanned by using a Affymetrix GeneTitan chip platform (Thermofisher, U.S.A.), a chip scanning result is obtained by using Axiomanalysisoxite_5.1.1 software to read fluorescent signals and convert the fluorescent signals into SNP, and the result shows that 96 individuals are successfully subjected to uniform typing, the typing success rate is 100%, and the typing accuracy of the chip is high.
3. Chip data filtering and analysis
And converting the SNP data into a Plink format (https:// zzz.bwh. Harvard.edu/Plink /), counting the minimum allele frequency of each SNP locus by using Plink software, filtering the SNP loci according to the typing success rate being more than 95%, and filtering the SNP loci according to the minimum allele frequency being more than 0.05, wherein the locus typing success rate of a single individual is more than 95% as a standard, and obtaining 394722 high-quality and high-polymorphism SNP loci. Statistical filtered SNP allele frequency information using R software (http:// www.r-project. Org /) is shown in FIG. 2. As can be seen from FIG. 2, the obtained SNP markers have high polymorphism and a relatively uniform distribution of MAF.
4. Genetic relationship analysis of different populations
The 394722 SNP data described above were imported into GCTA (Yang J, lee S H, goddard M E, et al GCTA: a tool for genome-wide complex trait analysis. The American Journal of Human Genetics,2011,88 (1): 76-82) to calculate the relationships between individuals, and the R-package Pheatmap (https:// giub. Com/raivokolde/ph) was used to map the relationship heat map, and ggplot2 (https:// github. Com/tidyverse/ggplot 2) was used to map the PCA between groups (FIG. 3). As can be seen from fig. 3, the individuals of the A, B, C population are clustered together, respectively, and there is a significant genetic distance between the three populations, so that different germplasm can be accurately distinguished by using chip data.
5. Genetic breeding analysis
The SNP data can be used for carrying out whole genome association analysis by using software such as GEMMA, rMVP and the like according to the phenotype data, and the SNP markers related to the traits can be positioned, and the HIBLUP, BGLR and the like can be used for carrying out whole genome selection analysis, so that accurate genetic evaluation of individuals is realized.
While the invention has been described in detail in terms of general description and specific embodiments, it will be apparent to those skilled in the art that modifications and improvements can be made to the scope of the invention. Accordingly, such modifications or improvements may be made without departing from the spirit of the invention and are intended to be within the scope of the invention as claimed.
Table 1: SNP locus information table of core No. 1 gene chip of Litopenaeus vannamei. The chromosome is the number of LVANscafold, the position is the position of SNP locus on scafold, and SNP is the mutation type of the locus.
Claims (6)
1. A litopenaeus vannamei whole genome SNP chip is characterized in that: the chip contains 594875 SNP sites, and the information of the SNP is shown in Table 1.
2. The method for preparing the whole genome SNP chip of the litopenaeus vannamei as set forth in claim 1, which is characterized in that:
(1) Screening out high-quality SNP loci according to MAF >0.05 by utilizing genome variation data of the litopenaeus vannamei, and constructing an SNP database of the litopenaeus vannamei;
(2) Analyzing the sequence around the SNP locus, deleting the SNP locus which cannot be designed by the probe, and deleting loci containing other SNPs within the range of 35bp around the SNP locus;
(3) On the genome, taking 2kb as a sliding window, randomly selecting 1 SNP locus in each window to form SNP data of genome even distribution;
(4) Screening SNP loci positioned in an exon region according to the annotation information of the SNP, adding the SNP loci into the obtained genome uniform distribution SNP data, and selecting 1 SNP locus in each window according to 500bp serving as a sliding window, so that the genome exon region; meanwhile, adding the SNP locus of the splice into the SNP data set;
(5) Adding the sites related to the characters into the SNP data set, preparing the SNP data set by using a chip technology, and finally obtaining the 600K SNP typing chip of the Litopenaeus vannamei genome, wherein the SNP typing chip contains 594875 SNP sites in total.
3. A method of preparation as claimed in claim 2, wherein: and (3) the neutral shape of the character related sites in the step (5) is growth, WSSV resistance, sex resistance, AHPND resistance and survival rate.
4. The use of the whole genome SNP chip of litopenaeus vannamei as set forth in claim 1, characterized in that: the chip is applied to germplasm genetic relationship analysis and germplasm identification of prawns.
5. The use of the whole genome SNP chip of litopenaeus vannamei as set forth in claim 1, characterized in that: the application of the Litopenaeus vannamei whole genome SNP chip in identification of associated genes of prawn characters.
6. The use of the whole genome SNP chip of litopenaeus vannamei as set forth in claim 1, characterized in that: the application of the Litopenaeus vannamei whole genome SNP chip in the whole genome selective breeding of the prawns.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310120282.8A CN116287297A (en) | 2023-02-16 | 2023-02-16 | Litopenaeus vannamei whole genome SNP chip and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310120282.8A CN116287297A (en) | 2023-02-16 | 2023-02-16 | Litopenaeus vannamei whole genome SNP chip and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116287297A true CN116287297A (en) | 2023-06-23 |
Family
ID=86778917
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310120282.8A Pending CN116287297A (en) | 2023-02-16 | 2023-02-16 | Litopenaeus vannamei whole genome SNP chip and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116287297A (en) |
-
2023
- 2023-02-16 CN CN202310120282.8A patent/CN116287297A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN113667760B (en) | SSR (simple sequence repeat) marker primer and method for evaluating genetic diversity of sparus praecox population | |
| CN115232880B (en) | Hainan black goat liquid-phase chip and application thereof | |
| CN115992265B (en) | Grouper whole genome liquid phase chip and application thereof | |
| CN102168136B (en) | Application of LHCGR gene of Chinese Holstein cow used as molecular marker | |
| CN116516029A (en) | Golden pomfret whole genome breeding chip and application | |
| CN109486961B (en) | Construction method of scylla paramamosain high-density genetic map | |
| Wang et al. | Development and evaluation of liquid SNP array for large yellow croaker (Larimichthys crocea) | |
| CN117487933A (en) | SSR (simple sequence repeat) marker primer, method and application for evaluating genetic diversity of puffer fish population | |
| Qin et al. | Transcriptomics analysis of cashmere fineness functional genes | |
| CN114875157B (en) | SNP (Single nucleotide polymorphism) marker related to individual growth traits of pelteobagrus fulvidraco and application | |
| CN116287297A (en) | Litopenaeus vannamei whole genome SNP chip and application thereof | |
| CN106701930B (en) | Method for detecting sheep FTH-1 gene insertion deletion polymorphism by using PCR-SSCP (polymerase chain reaction-single strand conformation polymorphism) and application thereof | |
| CN113604587A (en) | Molecular marker T5198 for rapidly identifying low-temperature tolerant variety of penaeus japonicus and application thereof | |
| CN113621714B (en) | Low-temperature-resistant molecular marker A257 of penaeus japonicus and application thereof | |
| Rajawat et al. | Genome-wide mining of diversity and evolutionary signatures revealed selective hotspots in Indian Sahiwal cattle | |
| CN117683911B (en) | Primer for detecting SNP molecular markers related to protein utilization characteristics of garrupa feed and application of primer | |
| CN114457168B (en) | Detection primer of microsatellite related to body color black spot character of red tilapia in wintering period and application of detection primer | |
| CN113897443B (en) | SNP molecular marker related to milk fat percentage of southern Holstein cows, kit and application and breeding method thereof | |
| CN116970712B (en) | SNP molecular marker related to reproduction traits on goat chromosome 13 and application | |
| CN116287296A (en) | SNP chip suitable for whole genome breeding application of litopenaeus vannamei and application | |
| CN108060236A (en) | A kind of method based on SNP site discriminating Jinhua Pigs, Large White | |
| CN110791511B (en) | Hypoxia-resistant megalobrama amblycephala growth character gene and positioning method and application thereof | |
| CN114300042A (en) | Method for screening candidate markers related to reproductive performance of white pigs based on whole genome correlation analysis | |
| CN116904607A (en) | SNP molecular marker combination for screening meat quality traits of Guan Ling cattle and application thereof | |
| CN117402980A (en) | Fugu rubripes genome-wide SNP locus combination and application thereof |
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 |