CN117126948B - Molecular marker combination for analyzing goat ear characters and application thereof - Google Patents
Molecular marker combination for analyzing goat ear characters and application thereof Download PDFInfo
- Publication number
- CN117126948B CN117126948B CN202311361134.1A CN202311361134A CN117126948B CN 117126948 B CN117126948 B CN 117126948B CN 202311361134 A CN202311361134 A CN 202311361134A CN 117126948 B CN117126948 B CN 117126948B
- Authority
- CN
- China
- Prior art keywords
- goat
- analyzing
- ear
- snp
- size
- 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.)
- Active
Links
- 241000283707 Capra Species 0.000 title claims abstract description 98
- 239000003147 molecular marker Substances 0.000 title abstract description 6
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 32
- 238000000034 method Methods 0.000 claims abstract description 24
- 239000003068 molecular probe Substances 0.000 claims abstract description 13
- 101100163849 Arabidopsis thaliana ARS1 gene Proteins 0.000 claims abstract description 4
- 101100097319 Schizosaccharomyces pombe (strain 972 / ATCC 24843) ala1 gene Proteins 0.000 claims abstract description 4
- 239000000523 sample Substances 0.000 claims description 25
- 210000005069 ears Anatomy 0.000 claims description 12
- 238000004458 analytical method Methods 0.000 claims description 10
- 238000012360 testing method Methods 0.000 claims description 4
- 238000002864 sequence alignment Methods 0.000 claims description 3
- 101100479039 Caenorhabditis elegans aars-1 gene Proteins 0.000 claims 1
- 238000009395 breeding Methods 0.000 abstract description 9
- 230000001488 breeding effect Effects 0.000 abstract description 9
- 230000002068 genetic effect Effects 0.000 abstract description 8
- 238000011156 evaluation Methods 0.000 abstract description 7
- 230000008569 process Effects 0.000 abstract description 6
- 238000004904 shortening Methods 0.000 abstract 1
- 238000001514 detection method Methods 0.000 description 12
- 108020004414 DNA Proteins 0.000 description 11
- 102000053602 DNA Human genes 0.000 description 11
- 238000007796 conventional method Methods 0.000 description 7
- 235000013372 meat Nutrition 0.000 description 7
- 210000002268 wool Anatomy 0.000 description 7
- 210000004080 milk Anatomy 0.000 description 5
- 239000008267 milk Substances 0.000 description 5
- 235000013336 milk Nutrition 0.000 description 5
- 239000002773 nucleotide Substances 0.000 description 5
- 125000003729 nucleotide group Chemical group 0.000 description 5
- 238000002360 preparation method Methods 0.000 description 5
- 238000012163 sequencing technique Methods 0.000 description 5
- 230000008901 benefit Effects 0.000 description 4
- 239000003153 chemical reaction reagent Substances 0.000 description 4
- 230000006872 improvement Effects 0.000 description 4
- 238000012216 screening Methods 0.000 description 4
- 108091028043 Nucleic acid sequence Proteins 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 102100025250 C-X-C motif chemokine 14 Human genes 0.000 description 2
- 244000025254 Cannabis sativa Species 0.000 description 2
- 102100030309 Homeobox protein Hox-A1 Human genes 0.000 description 2
- 101000858068 Homo sapiens C-X-C motif chemokine 14 Proteins 0.000 description 2
- 101001083156 Homo sapiens Homeobox protein Hox-A1 Proteins 0.000 description 2
- 101000651912 Homo sapiens Homeobox protein SIX2 Proteins 0.000 description 2
- 101001094737 Homo sapiens POU domain, class 4, transcription factor 3 Proteins 0.000 description 2
- 101000819088 Homo sapiens Transcription factor GATA-6 Proteins 0.000 description 2
- 101000891326 Homo sapiens Treacle protein Proteins 0.000 description 2
- 101150082088 MSRB3 gene Proteins 0.000 description 2
- 102100028720 Methionine-R-sulfoxide reductase B3 Human genes 0.000 description 2
- 102100035398 POU domain, class 4, transcription factor 3 Human genes 0.000 description 2
- 241001494479 Pecora Species 0.000 description 2
- 102100021382 Transcription factor GATA-6 Human genes 0.000 description 2
- 102100040421 Treacle protein Human genes 0.000 description 2
- 239000000872 buffer Substances 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 238000012217 deletion Methods 0.000 description 2
- 230000037430 deletion Effects 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000010586 diagram Methods 0.000 description 2
- 230000008030 elimination Effects 0.000 description 2
- 238000003379 elimination reaction Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 230000002349 favourable effect Effects 0.000 description 2
- 239000004459 forage Substances 0.000 description 2
- 239000011521 glass Substances 0.000 description 2
- 230000003100 immobilizing effect Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 238000005457 optimization Methods 0.000 description 2
- 210000005259 peripheral blood Anatomy 0.000 description 2
- 239000011886 peripheral blood Substances 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 108700028369 Alleles Proteins 0.000 description 1
- 238000000018 DNA microarray Methods 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 102100027332 Homeobox protein SIX2 Human genes 0.000 description 1
- 102000003960 Ligases Human genes 0.000 description 1
- 108090000364 Ligases Proteins 0.000 description 1
- 108091092919 Minisatellite Proteins 0.000 description 1
- 239000000020 Nitrocellulose Substances 0.000 description 1
- 239000004677 Nylon Substances 0.000 description 1
- 241000283903 Ovis aries Species 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- 238000012271 agricultural production Methods 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 210000000077 angora Anatomy 0.000 description 1
- 239000011324 bead Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 101150024923 da gene Proteins 0.000 description 1
- 230000008021 deposition Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000035558 fertility Effects 0.000 description 1
- 230000004907 flux Effects 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000012215 gene cloning Methods 0.000 description 1
- 210000004209 hair Anatomy 0.000 description 1
- 238000009396 hybridization Methods 0.000 description 1
- 238000003780 insertion Methods 0.000 description 1
- 230000037431 insertion Effects 0.000 description 1
- 230000006651 lactation Effects 0.000 description 1
- 239000010985 leather Substances 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 230000004807 localization Effects 0.000 description 1
- 230000036244 malformation Effects 0.000 description 1
- 238000013507 mapping Methods 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 229920001220 nitrocellulos Polymers 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 150000007523 nucleic acids Chemical group 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 229920001778 nylon Polymers 0.000 description 1
- 239000004033 plastic Substances 0.000 description 1
- 229920000307 polymer substrate Polymers 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 238000012253 re-sequencing analysis Methods 0.000 description 1
- 238000003753 real-time PCR Methods 0.000 description 1
- 230000003716 rejuvenation Effects 0.000 description 1
- 238000007894 restriction fragment length polymorphism technique Methods 0.000 description 1
- 231100000241 scar Toxicity 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 238000007493 shaping process Methods 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- 210000003491 skin Anatomy 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 230000007704 transition Effects 0.000 description 1
- 238000013519 translation Methods 0.000 description 1
- 230000014616 translation Effects 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
-
- 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
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 discloses a molecular marker combination for analyzing goat ear traits and application thereof, relates to the technical field of biology, and provides 1173 molecular markers capable of analyzing goat ear traits, wherein physical position information of the molecular markers is determined based on comparison of genome sequences of goat reference genome ARS1, and a molecular probe combination, a gene chip and a kit prepared from 1173 SNP molecular markers can be used for carrying out genetic evaluation on goat individuals, judging and selecting early-stage difficult-to-measure goat ear traits, shortening generation intervals and accelerating breeding processes, so that a large amount of breeding cost is saved.
Description
Technical Field
The invention relates to the technical field of biology, in particular to the technical field of biological detection, and more particularly relates to goat ear size SNP locus combination and application thereof.
Background
During the natural environment selection or artificial breeding process, the species can gradually form unique appearance morphology and body type characteristics. According to statistics, 69 goat breeds exist in China at present, and 58 goat breeds exist in China. According to the natural geographical region division of breeding, goats in China can be divided into three regions: comprises the north of a yellow river and a Wei river, the Qinghai-Tibet plateau and the north area thereof, a region: the goat is mainly in the form of wool and milk, the wool length is dense, the wool is more and thicker, and the region is favorable for developing the wool goat and the milk goat; two areas: the method comprises the steps of a region between a yellow river, a Wei river and a Yangtze river, wherein the climate of the region is neither high cold nor dry nor too high temperature and high humidity, and goats in the region have the characteristics of a first region and a second region at the same time, are mainly purified and rejuvenated, and have great potential for developing goat production for cultivating specialized goats; three regions: the sheep in the three areas mainly comprise meat and fur or skin, meat and milk in the regions of Yangtze river, the fur is short and thin, the rainwater in the south is sufficient, the climate is mild, the growth period of forage grass is long, the forage grass resources and agricultural and sideline products are rich, the development of varieties with meat, fur or skin, meat and milk is facilitated, and the local goats have the characteristics of high fertility and rapid growth of lambs. Goats can be generally classified into the following six classes according to production uses: firstly, a meat goat: the meat is delicious and has small smell of mutton, and the meat is taken as a main feeding target; secondly, goat for wool: the fineness and the count of the wool are different with the high wool yield as the main feeding target; thirdly, goat for down production: the fluff of Mao Naceng is the main product; fourthly, a leather goat: the main product is sheepskin; fifthly, a goat for milk: the main feeding goal is high lactation; and six, the goat is a common local goat. However, there were few studies on goat ears.
Ear type is an important trait of current livestock breeds, and ear size is one of its important phenotypes. The goat ears are not only one of important goat traits, but also popular food materials with high nutritional value and medicinal value and good taste; in addition, the ear character of the goat has important significance in the source tracing of the goat variety and the protection, development and utilization of germplasm resources.
Aiming at the current goats with various varieties and changeable characters, how to rapidly screen the goats and distinguish the types of the ears of the goats becomes an urgent problem to be solved, therefore, the design of the SNP chip which is applicable to the Chinese goats and can rapidly and effectively detect the characters of the ears has very important significance.
Disclosure of Invention
In order to meet the requirements of the current goat variety research and agricultural production in China on the detection of the goat ear traits, the invention provides a molecular probe combination, a gene chip, a kit and application for analyzing the goat ear traits, and the site information provided by the invention can be used for rapidly and accurately realizing the deposition evaluation of the goat ear, variety screening, variety identification, variety tracing and goat breeding, is favorable for germplasm resource protection and germplasm resource improvement, and has the advantages of short time consumption, low cost and wide market benefit.
In order to achieve the technical purpose of the invention, the invention provides the following technical scheme:
in a first aspect, the present invention provides an application of 1173 SNP site combinations in analyzing goat ear sizes, wherein physical positions of the 1173 SNP site combinations are shown in table 1:
table 1 1173 location information for a combination of sites
Its physical location was determined based on goat reference genome ARS1 genome sequence alignment.
A second aspect provides a method of analyzing the ear size of a goat, comparing 1173 SNP site genotypes of genomic DNA of a goat to be tested with the 1173 SNP site genotypes of genomic DNA of a control goat;
wherein the 1173 SNP sites are 1173 SNP sites as set forth in Table 1.
In a third aspect, a molecular probe set for analyzing the ear size of a goat is provided, wherein the molecular probe set detects SNP site combinations as shown in Table 1 in a sample to be tested, and the physical position information of the site combinations in Table 1 is determined based on the alignment of ARS1 genome sequences of a reference genome of the goat.
The fourth aspect is a gene chip for analyzing goat ear size, the gene chip being loaded with the molecular probe combination according to the third aspect.
The fifth aspect is a kit for analyzing goat ear size, which has the molecular probe set according to the third aspect or the gene chip according to the fourth aspect.
The sixth aspect is a method for analyzing the size of goat ears, which uses the molecular probe combination of the third aspect or the gene chip of the fourth aspect or the kit of the fifth aspect to detect a sample to be detected.
The molecular probe set according to the seventh aspect, the third aspect or the gene chip according to the fourth aspect or the kit according to the fifth aspect has any of the following uses:
(1) Application in goat ear size evaluation;
(2) Application in goat variety screening;
(3) Application in goat variety identification;
(4) Application in the tracing of goat breeds;
(5) Application in goat breeding;
(6) Application in germplasm resource protection;
(7) Application in germplasm resource improvement;
(8) The application in goat pedigree reconstruction.
Advantageous effects
1. The SNP locus combination provided by the invention has good domestic and foreign universality, can rapidly evaluate the early-period non-obvious goat ear size characters from the gene level, acquires more accurate breeding evaluation information, controls the breeding process, can screen, identify and trace the source of goat varieties by utilizing the locus combination, and provides technical support for germplasm resource protection and germplasm resource improvement.
2. The probe combination, the gene chip and the kit for analyzing the ear size of the goat provided by the invention have the characteristics of small flux, low cost and easier analysis, and have wide universality and wide market prospect.
Drawings
FIG. 1 is a Manhattan diagram of Pak-angora, kamori, daer, matebele Vs Toggenburg set;
fig. 2 is a graph of the results of the present application for performing a significance test on the determination results of population threshold analysis.
Detailed Description
The invention will be further elucidated with reference to the following detailed description of specific embodiments, which are merely illustrative and are not to be construed as limiting the invention. Unless otherwise indicated, the techniques used in the examples are conventional and well known to those skilled in the art, and may be carried out with reference to the "bioinformatics and functional genomics" original third edition or related books, and the bioinformatics software and products used are also commercially available. The various processes and methods not described in detail are conventional methods known in the art, the source of the materials used, the trade names and those necessary to list the constituents are all indicated at the first occurrence, and the same reagents used thereafter, unless otherwise indicated, are all the same as the first indicated.
In addition, it should be noted that the site combination and application provided by the invention are accomplished by the inventor through hard creative work and optimization work.
The features and advantages described in the previous site combination section herein are equally applicable to the combination of molecular probes formed based on site combination, gene chips, kits and uses thereof, and are not described in detail herein.
The size of the goat ear is divided according to the size of the ear area, the ear area of the conventional area is a big ear, the ear length is generally 25 to 35 cm, and the flat width of the ear is 12 to 15 cm; the malformation of the mutated small ear is the small ear.
The SNP referred to in the present invention means a single nucleotide polymorphism (Single Nucleotide Polymorphism), mainly means a DNA sequence polymorphism caused by a variation of a single nucleotide including a variation caused by a single base transition, a transversion, an insertion or a deletion at the genome level.
It should be noted that the molecular marker of the present invention is any heritable and detectable DNA sequence or protein, including, but not limited to, molecular markers based on molecular hybridization, such as RFLP, minisatelliteDNA; molecular markers based on PCR technology, such as RAPD, STS, SSR and SCAR; DNA markers based on restriction and PCR techniques; molecular markers based on DNA chip technology, such as SNPs; analytical marking techniques developed based on EST databases, and the like. The molecular marker provided by the invention can be used for genome mapping, gene localization research, gene cloning based on a map, species genetic relationship, system classification and the like.
The probe referred to in the present invention is a nucleic acid sequence (DNA or RNA) with a known sequence complementary to the target gene, such as Taqman-MGB probe.
It should be noted that the kit of the present invention is any one of the kits conventionally used in the art, which contains reagents used for detection or experiment, and is convenient for operators to get rid of the heavy reagent preparation and optimization process. In one embodiment of the invention, the kit comprises a primer for amplifying the site information provided by the invention, a molecular marker or a probe or a gene chip for detecting the site information provided by the invention, an enzyme and a buffer used for amplification, or a fluorescent marker used for detection.
Example 1 obtaining ear trait SNP site combinations
1. Selection of goat individuals
In order to realize more comprehensive coverage of domestic and foreign goat breeds, 361 individuals of 68 goat breeds in the global scope are re-sequenced, and each breed is:
the preparation method comprises the following steps of, menabe coat Mei Nabei Goat.
It should be noted that, some kinds of goats have no formal Chinese translations, and English names are still used at present.
2. Acquisition of Total SNP set of goat Whole Gene
The sample carrying genetic information from the individual goat in step 1 is collected using methods conventional in the art, including but not limited to blood, cells, tissue, skin, hair, excrement, etc. Genetic information (such as DNA) in the sample is extracted for high-depth sequencing, and comparison is carried out on a reference genome (obtained from NCBI) of a goat published in 2016 by using two modes of SAMtools and GATK, and a common result obtained by the comparison of the two modes forms a SNP set which contains 17,668,737 single nucleotide variations at the whole genome level. From these breeds, 36 goat individuals related to the ear size were selected.
The genetic information (genetic information) referred to herein refers to information that an organism is to replicate the same thing as itself, is transferred from a parent to a progeny, or is transferred from a cell to a cell each time each cell divides.
It should be noted that, the high-depth sequencing by extracting genetic information (e.g., DNA) in the sample may be performed by a biological company, such as hua da gene company, illuminea company, etc., and the high-depth sequencing method is a conventional method in the art or a method of the biological company, and in one embodiment of the present invention, the high-depth sequencing is performed by using an average sequencing depth of about 25.7×, and using a re-sequencing analysis procedure.
3. Screening of candidate genes and located functional regions
Grouping according to the remarkable differences of goat varieties in ear traits worldwide, summarizing all varieties of individuals with big ears and small ears, and performing selective elimination analysis according to the two groups of big ears and small ears. The selected Pak-angora (Pakistan angora), kamori, daer (Jianzhou Dart), matebile (Martebeli) and Toggenburg (ipenburg goat) small-ear sheep were grouped for elimination analysis.
The invention scans the functional area related to the ear size by calculating the nucleotide base polymorphism and locus allele frequency difference between pi value and fst value and scans the functional area related to the ear size by taking intersection of the calculation results of the two methods, screens out the functional area related to the ear size, screens out genes in the area by referring to the Manhattan diagram which is generated by different groups of ear size characters and is made by pi value and is shown in figure 1, and finally determines 7 candidate genes SIX2, CXCL14, POU4F3, TCOF1, HOXA1, MSRB3 and GATA6 which are related to the ear characters and have very definite functions. And then determining the functional region corresponding to the candidate gene by perl script.
4. Ear-sized SNP site combination acquisition
Searching SNP loci corresponding to the functional regions of the candidate genes determined in the step 3 in the total SNP set by utilizing bedtools to obtain ear trait locus combinations which are related to 7 ear traits in total and only comprise 1173 SNP loci, wherein the functional genes are related to SIX2, CXCL14, POU4F3, TCOF1, HOXA1, MSRB3 and GATA6.
Example 2 ear trait SNP locus combination for preparation of primer combination and Probe combination
The technical personnel designs the primer according to the sequence information of each site in the ear character SNP site combination provided by the invention, and carries out secondary structure evaluation and Tm value evaluation on the designed primer, so that the primer with good specificity and high sensitivity can be finally obtained, and the detection purpose can be realized under the same reaction condition.
The secondary structure and Tm value can be evaluated by any means commonly used in the art, for example, by using DNA shaping Form to evaluate the secondary structure, see (http:// unfold. Rna. Albany. Edu// q=mfold/DNA-shaping-Form), and then using software RaW-Probe to evaluate the Tm value.
The method is a conventional method, and the site information in the SNP site combination with the ear size can be obtained without creative labor, so that the primer obtained by the SNP site combination provided by the invention also belongs to the protection scope of the invention.
Likewise, probes prepared by using the SNP site combinations provided by the invention, such as tanqman probes, also belong to the scope of the invention.
Example 3 SNP site combination for analysis of ear size for preparation of Gene chip
The SNP gene chip of the invention is obtained by immobilizing the primer or probe obtained in example 2 on a polymer substrate such as nylon membrane, nitrocellulose membrane, plastic, silica gel wafer, micro magnetic beads or the like by a conventional method, or immobilizing the probe on a glass plate, or directly synthesizing the primer or probe obtained in example 2 on a hard surface such as glass, and the method of using the SNP gene chip of the invention is the same as that of the conventional method.
It should be noted that, a person skilled in the art can prepare an SNP gene chip for detecting the ear size of a goat in any one of the ways, and can also entrust the preparation of biological companies, but all SNP gene chips prepared based on the combination of SNP loci for ear size analysis provided by the application belong to the protection scope of the invention.
Example 4 analysis kit for goat ear traits
The SNP detection kit for ear size analysis provided by the application comprises a primer or a probe or a gene chip obtained based on the SNP locus combination obtained in example 1. Depending on the type of use, corresponding detection reagents are also included, such as buffers, ligases, aceQUniversal u+ Probe Master Mix V2, taqman probes, etc., conventionally used for fluorescent quantitative PCR reactions when Taqman probes are obtained based on the SNP site combinations obtained in example 1.
Different SNP kits for detecting the ear traits of the goat are configured by a person skilled in the art according to different using modes, but the SNP detection kits for detecting the ear traits of the goat based on the ear trait SNP locus combination configuration provided by the application belong to the protection scope of the invention.
Example 5 detection of goat ear Properties
Based on the SNP locus combination for analyzing the ear size of the goat provided in the embodiment 1 of the application, the ear size of the goat with known ear size is detected, and according to the detection result, the accuracy of detection is judged by combining with the known ear size, specifically:
collecting peripheral blood of goats by adopting a conventional method, and extracting whole genome DNA (deoxyribonucleic acid) in the peripheral blood to obtain a whole genome DNA sample;
the gene chip is designed according to the site information in the SNP site combination provided by the invention by adopting a conventional method, a whole genome DNA sample of the goat is detected, a typing result of each site in the goat (namely, whether each site is homozygote, heterozygote, mutant homozygote or base deletion result) is obtained, the frequency value of the typing result of each site is calculated, the frequency value is compared with a population threshold value, and the comparison result shows that the gene detection result is consistent with the corresponding goat ear phenotype.
The population threshold value is obtained by analyzing populations with different ear sizes, and the method is the same as the above.
The method has the advantages that the significance test (independent sample Manwhitney U test) is carried out on the judging result of the analysis of the large-ear goat population and the small-ear goat population, the result is shown in figure 2, according to the result in the figure, P is less than 0.01, the difference is extremely significant, and the accuracy and the effectiveness of the judging result by adopting the method are realized.
Industrial application
The SNP locus combination which is only composed of 1173 SNP loci and can be used for analyzing the ear size of a goat and can be prepared by a person skilled in the art, a gene chip and a kit can analyze the ear size of a goat individual from genome level, genetic information evaluation, variety screening and variety identification can be realized, the breeding process can be controlled, and the SNP locus combination can be applied to goat variety tracing, goat pedigree reconstruction, germplasm resource protection and germplasm resource improvement.
The above description is only for the purpose of aiding in understanding the preferred embodiments of the present invention and is not intended to limit the present invention, and various changes and modifications may be made to the present invention by those skilled in the art without departing from the spirit of the present invention, and it should also be considered that the present invention shall fall within the scope of the present invention.
Claims (6)
- Application of 1.1173 SNP locus combinations in analysis of goat ear sizes, physical positions of 1173 SNP locus combinations are shown in the following table:；its physical location is determined based on genomic sequence alignment of goat reference genome ARS 1.
- 2. A method for analyzing the ear size of a goat, wherein 1173 SNP locus genotypes of genomic DNA of the goat to be detected are compared with 1173 SNP locus genotypes of genomic DNA of a control goat;wherein the 1173 SNP sites are 1173 SNP sites of claim 1.
- 3. Analyzing a goat ear size molecular probe set which detects the SNP site set as set forth in claim 1 in a test sample, the physical location information of the site set in claim 1 being determined based on goat reference genome ARS1 genome sequence alignment.
- 4. A gene chip for analyzing the size of goat ears, which is loaded with the molecular probe combination of claim 3.
- 5. A kit for analyzing the size of goat ears, which has the molecular probe set according to claim 3 or the gene chip according to claim 4.
- 6. A method for analyzing the size of goat ears, which comprises detecting a sample to be detected using the molecular probe set of claim 3 or the gene chip of claim 4 or the kit of claim 5.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311361134.1A CN117126948B (en) | 2023-10-20 | 2023-10-20 | Molecular marker combination for analyzing goat ear characters and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311361134.1A CN117126948B (en) | 2023-10-20 | 2023-10-20 | Molecular marker combination for analyzing goat ear characters and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN117126948A CN117126948A (en) | 2023-11-28 |
| CN117126948B true CN117126948B (en) | 2024-02-09 |
Family
ID=88860325
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311361134.1A Active CN117126948B (en) | 2023-10-20 | 2023-10-20 | Molecular marker combination for analyzing goat ear characters and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117126948B (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR102243308B1 (en) * | 2019-11-08 | 2021-04-22 | 대한민국 | Novel SNP marker for identification of black goat and uses thereof |
| CN113265475A (en) * | 2021-07-21 | 2021-08-17 | 中国农业大学 | Gene chip for analyzing sheep fat tail, molecular probe combination, kit and application |
| CN113293220A (en) * | 2021-07-23 | 2021-08-24 | 中国农业大学 | Gene chip for analyzing ear size of sheep, molecular probe combination, kit and application |
| CN113396866A (en) * | 2021-05-28 | 2021-09-17 | 河南省畜牧总站 | Method for breeding new variety of meat goats |
-
2023
- 2023-10-20 CN CN202311361134.1A patent/CN117126948B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR102243308B1 (en) * | 2019-11-08 | 2021-04-22 | 대한민국 | Novel SNP marker for identification of black goat and uses thereof |
| CN113396866A (en) * | 2021-05-28 | 2021-09-17 | 河南省畜牧总站 | Method for breeding new variety of meat goats |
| CN113265475A (en) * | 2021-07-21 | 2021-08-17 | 中国农业大学 | Gene chip for analyzing sheep fat tail, molecular probe combination, kit and application |
| CN113293220A (en) * | 2021-07-23 | 2021-08-24 | 中国农业大学 | Gene chip for analyzing ear size of sheep, molecular probe combination, kit and application |
Non-Patent Citations (2)
| Title |
|---|
| 利用混池测序初步筛选藏山羊耳面积性状的候选基因;王泽稷;中国优秀硕士学位论文全文数据库（电子期刊）(第1期);第D050-463页 * |
| 比较基因组学分析鉴定影响藏山羊耳朵大小的遗传位点;钟杰，等;西北农业学报;第28卷(第11期);第1742-1749页, 摘要，第1743页左栏，材料与方法, 第1746页，图4-5 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN117126948A (en) | 2023-11-28 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN113278712B (en) | Gene chip, molecular probe combination, kit and application for analyzing sheep hair color | |
| CN107619857B (en) | Method for detecting CNV (CNV) marker of beef cattle KLF8 gene and application of CNV marker | |
| CN113278716B (en) | Gene chip for analyzing characters for sheep wool, molecular probe combination, kit and application | |
| CN113265476B (en) | Gene chip, molecular probe combination, kit and application for analyzing milk production performance of sheep | |
| CN105506111B (en) | Method for detecting CNV (CNV) marker of MAPK10 gene of Nanyang cattle and application of CNV marker | |
| CN115029451B (en) | Sheep liquid phase chip and application thereof | |
| CN114134236B (en) | Application of reagent for detecting SNP molecular markers in goat RBP4 genotyping and/or goat molecular marker assisted breeding | |
| WO2023001209A1 (en) | Gene chip, molecular probe combination, test kit and application for analyzing sheep fat tails | |
| Salisu et al. | Molecular markers and their Potentials in Animal Breeding and Genetics | |
| CN114959059A (en) | SNP locus combination related to diameter variation coefficient of fine wool sheep wool fiber and application thereof | |
| CN113293220B (en) | Gene chip for analyzing ear size of sheep, molecular probe combination, kit and application | |
| CN113278714B (en) | Gene chip for analyzing whether sheep has horns or not, molecular probe combination, kit and application | |
| CN117126948B (en) | Molecular marker combination for analyzing goat ear characters and application thereof | |
| CN104769133A (en) | Method of improving microarray performance by strand elimination | |
| AU2023280943A1 (en) | Snp locus combinations related to diameter of fine wool sheep wool fibers, and use | |
| CN117089633B (en) | Molecular marker combination for analyzing existence of goat fluff and application | |
| CN117089635B (en) | Molecular marker combination for analyzing goat reproductive performance and application | |
| CN117089634B (en) | Molecular marker combination for analyzing goat milk performance and application | |
| CN117089636B (en) | Molecular marker combination for analyzing goat meat performance and application | |
| CN117106936B (en) | Molecular marker combination for analyzing goat hair color character and application | |
| CN114292924A (en) | Sika deer whole genome SNP molecular marker combination, SNP chip and application | |
| CN109811062B (en) | FSHR gene specific SNP marker, detection method of Tian Qiaoda lambing number character of red sheep and application of detection method | |
| CN113278713B (en) | Gene chip, molecular probe combination, kit and application of sheep multi-angle character | |
| CN117106935A (en) | Molecular marker combination for analyzing angular character of goat and application thereof | |
| CN113278715A (en) | Gene chip for analyzing sheep papilla number, molecular probe combination, kit and application |
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 | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |