CN116606938A - SNP locus related to improving innate immunity of yellow-feather broilers and application thereof - Google Patents
SNP locus related to improving innate immunity of yellow-feather broilers and application thereof Download PDFInfo
- Publication number
- CN116606938A CN116606938A CN202310607006.4A CN202310607006A CN116606938A CN 116606938 A CN116606938 A CN 116606938A CN 202310607006 A CN202310607006 A CN 202310607006A CN 116606938 A CN116606938 A CN 116606938A
- Authority
- CN
- China
- Prior art keywords
- snp
- disease resistance
- chicken
- chickens
- marker
- 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
- 241000287828 Gallus gallus Species 0.000 title claims abstract description 110
- 230000015788 innate immune response Effects 0.000 title abstract description 7
- 235000013330 chicken meat Nutrition 0.000 claims abstract description 89
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 40
- 210000002540 macrophage Anatomy 0.000 claims abstract description 35
- 208000035240 Disease Resistance Diseases 0.000 claims abstract description 30
- 238000000034 method Methods 0.000 claims abstract description 15
- 238000012216 screening Methods 0.000 claims abstract description 14
- 238000004458 analytical method Methods 0.000 claims abstract description 12
- 210000005259 peripheral blood Anatomy 0.000 claims description 20
- 239000011886 peripheral blood Substances 0.000 claims description 20
- 239000003550 marker Substances 0.000 claims description 16
- 239000003795 chemical substances by application Substances 0.000 claims description 8
- 210000000349 chromosome Anatomy 0.000 claims description 7
- 238000012163 sequencing technique Methods 0.000 claims description 7
- 238000003205 genotyping method Methods 0.000 claims description 4
- 239000002773 nucleotide Substances 0.000 claims description 4
- 125000003729 nucleotide group Chemical group 0.000 claims description 4
- 230000008030 elimination Effects 0.000 claims description 2
- 238000003379 elimination reaction Methods 0.000 claims description 2
- 229940088592 immunologic factor Drugs 0.000 claims description 2
- 108020004999 messenger RNA Proteins 0.000 claims description 2
- 210000002966 serum Anatomy 0.000 claims description 2
- 230000000242 pagocytic effect Effects 0.000 abstract description 23
- 108700028369 Alleles Proteins 0.000 abstract description 6
- 210000000988 bone and bone Anatomy 0.000 abstract description 6
- 230000008569 process Effects 0.000 abstract description 3
- 238000012248 genetic selection Methods 0.000 abstract description 2
- 239000000463 material Substances 0.000 abstract description 2
- 230000013011 mating Effects 0.000 abstract description 2
- 238000000926 separation method Methods 0.000 abstract description 2
- 210000003746 feather Anatomy 0.000 abstract 1
- 230000035772 mutation Effects 0.000 description 13
- 206010039438 Salmonella Infections Diseases 0.000 description 12
- 238000009395 breeding Methods 0.000 description 12
- 230000001488 breeding effect Effects 0.000 description 12
- 206010039447 salmonellosis Diseases 0.000 description 12
- 238000005516 engineering process Methods 0.000 description 6
- 230000036039 immunity Effects 0.000 description 6
- 210000004369 blood Anatomy 0.000 description 4
- 239000008280 blood Substances 0.000 description 4
- 244000144977 poultry Species 0.000 description 4
- 235000013594 poultry meat Nutrition 0.000 description 4
- 201000010099 disease Diseases 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- 210000003979 eosinophil Anatomy 0.000 description 3
- 210000004698 lymphocyte Anatomy 0.000 description 3
- 230000001717 pathogenic effect Effects 0.000 description 3
- 230000001105 regulatory effect Effects 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 102100032386 1,5-anhydro-D-fructose reductase Human genes 0.000 description 2
- 101150071667 H2AFZ gene Proteins 0.000 description 2
- 101000797917 Homo sapiens 1,5-anhydro-D-fructose reductase Proteins 0.000 description 2
- 101000775437 Homo sapiens All-trans-retinol dehydrogenase [NAD(+)] ADH4 Proteins 0.000 description 2
- 206010057249 Phagocytosis Diseases 0.000 description 2
- 230000010799 Receptor Interactions Effects 0.000 description 2
- 241000607142 Salmonella Species 0.000 description 2
- 230000006907 apoptotic process Effects 0.000 description 2
- 230000022131 cell cycle Effects 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 230000004069 differentiation Effects 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- 230000003993 interaction Effects 0.000 description 2
- 235000013372 meat Nutrition 0.000 description 2
- 230000037353 metabolic pathway Effects 0.000 description 2
- 230000010627 oxidative phosphorylation Effects 0.000 description 2
- 230000008782 phagocytosis Effects 0.000 description 2
- 210000000680 phagosome Anatomy 0.000 description 2
- 238000010187 selection method Methods 0.000 description 2
- 230000019491 signal transduction Effects 0.000 description 2
- MFYSUUPKMDJYPF-UHFFFAOYSA-N 2-[(4-methyl-2-nitrophenyl)diazenyl]-3-oxo-n-phenylbutanamide Chemical compound C=1C=CC=CC=1NC(=O)C(C(=O)C)N=NC1=CC=C(C)C=C1[N+]([O-])=O MFYSUUPKMDJYPF-UHFFFAOYSA-N 0.000 description 1
- 101150090724 3 gene Proteins 0.000 description 1
- 102100031795 All-trans-retinol dehydrogenase [NAD(+)] ADH4 Human genes 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 102100021287 CUE domain-containing protein 2 Human genes 0.000 description 1
- 108090000695 Cytokines Proteins 0.000 description 1
- 102000004127 Cytokines Human genes 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 241000223924 Eimeria Species 0.000 description 1
- 101800001649 Heparin-binding EGF-like growth factor Proteins 0.000 description 1
- 102100039385 Histone deacetylase 11 Human genes 0.000 description 1
- 108700038332 Histone deacetylase 11 Proteins 0.000 description 1
- 101000894806 Homo sapiens CUE domain-containing protein 2 Proteins 0.000 description 1
- 101000905054 Homo sapiens Histone H2A.Z Proteins 0.000 description 1
- 101000639792 Homo sapiens U2 small nuclear ribonucleoprotein A' Proteins 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N Phenol Chemical compound OC1=CC=CC=C1 ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 1
- 102100033762 Proheparin-binding EGF-like growth factor Human genes 0.000 description 1
- 102000058242 S100A12 Human genes 0.000 description 1
- 108700016890 S100A12 Proteins 0.000 description 1
- 101150097337 S100A12 gene Proteins 0.000 description 1
- 241001122767 Theaceae Species 0.000 description 1
- 102100034465 U2 small nuclear ribonucleoprotein A' Human genes 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 238000000246 agarose gel electrophoresis Methods 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 244000144974 aquaculture Species 0.000 description 1
- 238000009360 aquaculture Methods 0.000 description 1
- 238000009412 basement excavation Methods 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 238000010219 correlation analysis Methods 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 238000010201 enrichment analysis Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 244000144992 flock Species 0.000 description 1
- 238000000684 flow cytometry Methods 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000017448 oviposition Effects 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 230000004962 physiological condition Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000000638 stimulation Effects 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 238000011222 transcriptome analysis Methods 0.000 description 1
- 238000000870 ultraviolet spectroscopy Methods 0.000 description 1
- 210000003462 vein Anatomy 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
- 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)
- Life Sciences & Earth Sciences (AREA)
- Analytical Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Microbiology (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Biotechnology (AREA)
- Biophysics (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The invention provides SNP loci related to improving the innate immunity of yellow feather broilers and application thereof, local special chicken species, wenchang chickens and large bone chickens which are distributed in different natural geographic environments are taken as materials, disease resistance genes related to phagocytic functions of macrophages are identified by utilizing combined analysis of two varieties of genome and macrophage transcriptome data, SNP markers related to disease resistance of the broilers are obtained through screening, high disease resistance performance selection is realized through a screening related SNP frequency method, data support is provided for realizing accurate selection of characters with complicated disease resistance shapes and rapid homozygosity of character related alleles, genetic selection progress is accelerated, dominant allele loci of cultivated strains are in high frequency or pure and state, and the problem of offspring character separation can be avoided to a large extent in the process of applying new strains to mating system creation.
Description
Technical Field
The invention relates to the field of biology, relates to SNP loci related to improving innate immunity of yellow-feather broilers and application thereof, in particular to a method for culturing new-strain chickens with strong innate immunity by integrating and utilizing genome selection methods of SNP gene frequencies on genes with obvious differential innate immunity characteristics, belonging to the field of chicken innate immunity character improvement molecular breeding.
Background
The number of the chickens in China is over 100 hundred million in the world, and in recent years, along with the change of meat consumption structures in China, the consumption amount of chicken is in a trend of increasing year by year, and the chickens become the second largest meat consumption in China. However, the frequency of diseases is increased to cause serious economic loss to the broiler industry, and the excavation of disease-resistant genes and the cultivation of new varieties become the problems to be solved urgently. Local varieties such as Tibetan chickens, large-bone chickens, beijing oiled chickens and the like in China respectively have excellent characteristics of plateau adaptability, cold resistance, stronger disease resistance and the like; local chicken species such as Tibetan chickens, large mountain micro chickens, tea chickens and the like have strong resistance to eimeria and clear and distant spicy chickens have strong resistance to salmonella. So far, due to the limitation of the technology level of the original country of the variety in the field, the excellent genetic resources cannot be systematically evaluated and mined, in the prior broiler selection process, the attention of breeding workers is often focused on the growth characteristics, and the growth characteristics and the immune characteristics are often not directly related in the actual production. The phagocytic capacity of macrophages in peripheral blood is an important index for measuring the immunity of chicken species, is an important index for measuring the resistance of different chicken species to salmonella, and the strength of the phagocytic capacity directly influences the immunity of offspring and even the economic benefit of broiler chicken breeding.
In poultry breeding, the traditional method is difficult to improve individual immunity performance, meuwissen et al put forward (GS) in 2001, namely, the breeding value obtained by the method is called Genome Estimation Breeding Value (GEBV) by estimating all SNP marker effect values in the whole genome, the GS technology is widely applied to cattle, chickens, sheep and aquaculture, the GS technology can be used for broiler breeding due to the fact that a large number of available SNP numbers exist in chicken genome, the GS technology is applied to poultry breeding by the existing poultry breeding company internationally, various reports show that the GS technology is applied to selection of complex economic traits, the genotype of an individual is detected in a large amount by combining phenotype information, the whole genome correlation analysis and transcriptome sequencing technology, and candidate genes related to the target traits are found by SNP chips and transcriptome sequencing, so that the breeding cost can be reduced, the selection period can be shortened, and the selection accuracy can be improved.
Differences in immune traits among individuals are expressed in the ratio of peripheral blood eosinophils to lymphocytes (H/L) before and after salmonella infection, as well as the percentage of macrophages. Therefore, the blood related immunity index can be used as an index for measuring the immunity performance of broiler chickens to guide seed reserving. Under the large background of the rapid rise of the current poultry raising cost (feed, labor, environmental control and the like), the disease resistance of the broiler chickens is selectively improved by utilizing a genome and transcriptome selection method integrated with SNP, so that the cost of breeding new lines can be saved, the survival rate of individuals is improved, the breeding efficiency is improved by assistance, and the economic loss caused by frequent diseases to the broiler chickens industry is reduced.
Disclosure of Invention
In order to solve the technical problems, the invention aims at:
first, the present invention provides a screening method for an SNP on an integrated disease resistance candidate gene, comprising the steps of:
(1) Constructing a reference population and determining serum immune factor levels of both populations;
(2) Each chicken is sampled and preserved, DNA is extracted, and the SNP chip of Beijing core number one is utilized to carry out parting comprising the above 5 SNPs sites, and the data after genotyping is processed and the quality is controlled;
(3) Extracting peripheral blood macrophages of a 28-day-old uninfected Wenchang chicken (n=8) and a big-bone chicken (n=8) for transcriptome sequencing analysis;
(4) Transcriptome combined selection signal clearing analysis screens SNP loci which are obviously differentiated on candidate genes related to disease resistance.
Further, the invention provides a group of significant SNP markers which are related to the disease resistance of broiler chickens and located on genes of chicken chromosome 4 and influence disease resistance, and the detailed information is shown in the following table 1.
TABLE 1 significant SNP markers on genes affecting disease resistance traits
Further, the SNP marker specifically includes: C/T mutation at position 60,155,379 of chicken chromosome 4 (GGAZ), A/G at position 60,155,401 of chicken chromosome 4 (GGAZ) (reference genome: galGal6,), G/A mutation at position 60,155,459, A/G mutation at position 60,155,532, T/C mutation at position 60,155,544, G/T mutation at position 60,155,602, A/G mutation at position 60,155,886.
Further, the present invention provides a method for integrating SNP loci with significantly differentiated disease resistance traits and selecting a clone with higher disease resistance, comprising: obtaining genomic DNA from chicken and SNP chip-based whole genome SNPs; peripheral blood macrophage mRNA expression, screening candidate genes related to disease resistance by utilizing transcriptome expression difference and selective signal elimination analysis, screening out SNP (single nucleotide polymorphism) which is obviously differentiated on the candidate genes, and selecting individuals with high SNP gene frequency for seed reservation.
Advantageous effects
The research uses local special chicken species, wenchang chicken and big bone chicken which are distributed in different natural geographic environments as materials, utilizes the combined analysis of two varieties of genome and macrophage transcriptome data to identify disease resistance genes related to macrophage phagocytic function, screens and obtains SNP markers related to disease resistance of broiler chickens, realizes the selection of high disease resistance by screening related SNP frequency methods, provides data support for the accurate selection of characteristics with more complicated disease resistance shapes and the rapid homozygosity of characteristics related alleles, accelerates the genetic selection progress, ensures that dominant allele loci of cultivated strains are in high frequency or pure and state, and can also avoid the problem of offspring character separation to a large extent in the process of applying new strains to mating system creation.
Drawings
FIG. 1 ratio of peripheral blood eosinophils to lymphocytes (H/L) in large-bone chickens and Wenchang chickens before and after salmonella infection at 14 and 28 days of age.
FIG. 2 percentage of macrophages before and after Salmonella infection in both 14 and 28 day old large and Wenchang chickens.
Fig. 3 comparison of peripheral blood macrophage phagocytic capacity difference between 14-day-old and 28-day-old salmonella infection in large-bone chickens and Wenchang chickens (n=10 to 15).
FIG. 4 shows the correlation of the phagocytic index and the phagocytic rate of peripheral blood macrophages of 28-day-old Wenchang chickens and large-bone chickens.
FIG. 5 Wenchang and big bone chicken genome data population differentiation index (Fst) and nucleotide polymorphism (Pi) analysis.
Detailed Description
The terms used in the present invention generally have meanings commonly understood by those of ordinary skill in the art unless otherwise indicated. The invention will be described in further detail below in connection with specific examples and with reference to the data. It is to be understood that these examples are merely illustrative of the present invention and are not to be construed as limiting the scope of the invention, since numerous insubstantial modifications and adaptations of the invention will be apparent to those skilled in the art in light of the above disclosure.
Example 1 screening of SNPs on candidate genes for integration of disease resistance traits
(1) Determination of Experimental animals and target Properties
Local special chicken breeds, wenchang chicken and big bone chicken (see table 2) distributed in different natural geographic environments are selected as test groups, and 100 breeds are used for each breed. Salmonella infection was performed at 14 and 28 days of age, respectively, and the phagocytic capacity of macrophages in peripheral blood before and after salmonella infection was compared with those of Wenchang and Dagu chickens. And the disease resistance genes related to the phagocytic function of the macrophages are identified by utilizing the combined analysis of the genome data of the two varieties and the macrophage transcriptome data.
TABLE 2 descriptive statistics of H/L for 54 chickens
(2) Blood related immune index difference comparison of Wenchang chicken with big bones
The ratio of peripheral blood eosinophils to lymphocytes (H/L), and the percentage of macrophages, before and after salmonella infection, was counted for both 14 and 28 day old in large-bone chickens and Wenchang chickens. The results showed that the H/L value of peripheral blood of the large-bone chickens was significantly higher than that of Wenchang chickens, the H/L value was significantly increased after salmonella infection, and the H/L value was decreased with the increase of day age (FIG. 1). The H/L value difference between the big-bone chicken and Wenchang chicken is more remarkable at 28 days of age compared with 14 days of age. The peripheral blood macrophage percentage was more pronounced for the 14 day old and 28 day old large-bone chickens than for Wenchang chickens, with the 28 day old difference (FIG. 2).
(3) Macrophage phagocytic capacity comparison
Peripheral blood macrophages are extracted before and after salmonella infection, and phagocytosis index and phagocytosis rate of the macrophages are detected by adopting a flow cytometry. The results show (fig. 3) that, in the case of no infection, the phagocytic index and the phagocytic rate of macrophages of the large-bone chickens are higher than those of Wenchang chickens, but the phagocytic capacity of macrophages of the large-bone chickens after salmonella infection is obviously higher than those of Wenchang chickens without reaching a significant difference level. And at 28 days of age, when the chicken is not infected, the phagocytic capacity of macrophages of the large-bone chicken is obviously higher than that of Wenchang chicken, and after salmonella infection, the difference between the two is not obvious. The results show that the phagocytic capacity of the peripheral blood macrophages of the large-bone chickens is better than that of Wenchang chickens. The difference is not obvious in the early young and normal physiological conditions, but when the chicken is infected by a pathogen, the big bone chicken is more sensitive to pathogenic stimulus relative to Wenchang chicken peripheral blood macrophages, and the difference is caused to be more obvious. With the increase of day age, the immunity of the organism tends to be stable, and the phagocytic capacity of the peripheral blood macrophages of the large-bone chickens is still obviously higher than that of Wenchang chickens by 28 days of age, and the difference between the two chickens is reduced after salmonella infection. The result is synthesized that the phagocytic capacity of the macrophages of the peripheral blood of the large-bone chicken is stronger than that of Wenchang chicken, and the macrophages of the large-bone chicken at 14 days of age are more sensitive to pathogenic stimulus.
(4) Peripheral blood macrophage transcriptome analysis
Transcriptome sequencing was performed by extracting peripheral blood macrophages of 28-day-old Wenchang chickens (n=8) and large-bone chickens (n=8), respectively. The Fold Change is more than or equal to 1.5 and FDR is less than 0.05 as a screening standard, 1136 differential expression genes are screened altogether, wherein Wenchang chickens have 436 up-regulated genes and 700 down-regulated genes relative to large-bone chickens. These genes are involved mainly in signaling pathways such as cell cycle (cell cycle), cytokine interactions (cytokine-cytokine receptor interaction), ECM receptor interactions (ECM-receptor interaction), apoptosis (apoptosis), and phagosome (phagosome).
The correlation of phagocytic index and phagocytic rate of 98 samples was counted, and both showed a significant positive correlation (r2= 0.7733). WGCNA analysis was performed on macrophage phagocytic index and phagocytic rate phenotype, co-screening for three significantly related modules, the dark 2, ivory and lightyellow modules, respectively. Pathway enrichment analysis was performed on three modular genes, respectively, together into the metabolic pathway (metabolic pathways) and oxidative phosphorylation (oxidative phosphorylation) signaling pathways (fig. 4).
(5) Finding core genes in three modules (Hubgenes)
584 core genes are identified by taking the value of |KME|more than or equal to 8 as a threshold value, wherein 22 genes which are differentially expressed are highly expressed in Wenchang chickens. Gene functional annotation it was found that 6 genes were associated with cellular immune function, including CUEDC2, AKR1E2, ADH4, HBEGF, S100A12 and SNRPA1, where the AKR1E2 and ADH4 genes were significantly down-regulated after LPS stimulation of HD11 macrophages.
Macrophage candidate gene H2AFZ was found to differentiate significantly in Wenchang and large-bone chickens by the group differentiation index (Fst) and nucleotide polymorphism (Pi) of the Wenchang and large-bone chicken genome data (n=30). And the expression level of the gene in Wenchang chicken and large-bone chicken is obviously inversely related to the phagocytic index and the phagocytic rate of macrophages. A total of 24 mutation sites on the H2AFZ gene were polymorphic in medium angstrom breeder chickens (14 breeder chickens) and 679 individuals were subjected to polymorphism analysis, so that the gene frequencies of the sites in medium angstrom breeder chickens were polymorphic (FIG. 5).
The set of obvious SNP markers which are related to the disease resistance of the broiler chicken and are positioned on genes affecting the disease resistance of the chromosome 4 are obtained through screening by adopting a Ct value comparison method, and detailed information is shown in the following table.
The SNP marker specifically comprises: C/T mutation at position 60,155,379 of chicken chromosome 4 (GGAZ), A/G at position 60,155,401 of chicken chromosome 4 (GGAZ) (reference genome: galGal 6), G/A mutation at position 60,155,459, A/G mutation at position 60,155,532, T/C mutation at position 60,155,544, G/T mutation at position 60,155,602, A/G mutation at position 60,155,886.
Example 2 application of SNP on candidate Gene integrating disease resistance traits
1. Population to be selected
Chickens to be detected are randomly selected. The wing vein blood is collected after 280 days of age, ACD is anticoagulated, and the blood is preserved at-20 ℃ for standby.
2. DNA extraction
Genomic DNA was extracted by a conventional phenol-based method, dissolved in TE, and double-detected for purity and concentration by agarose gel electrophoresis and UV spectrophotometry, followed by dilution to a concentration of 50 ng/. Mu.l.
3. Genotyping
And sending and measuring chicken genome SNP chips, and determining genotypes of the SNP loci. The results of each chicken in the reference group were determined, and then the gene frequencies of SNPs on candidate genes were examined, and the differentiating SNPs on candidate genes in the control group and the test group are shown in Table 3.
TABLE 3 Gene frequencies of SNPs
| Site(s) | Position of | Control group | Test group |
| rs737893701 | 60155379 | 0.83 | 0.61 |
| rs735176537 | 60155401 | 0.90 | 0.62 |
| rs736904000 | 60155459 | 0.86 | 0.28 |
| rs738535625 | 60155532 | 0.80 | 0.23 |
| rs80690045 | 60155544 | 0.90 | 0.64 |
| rs80765089 | 60155602 | 0.85 | 0.62 |
| rs736133911 | 60155886 | 0.88 | 0.23 |
Allele detection of the amplified product is performed using direct sequencing or other effective means. Selecting and reserving according to genotyping results: because the selected sites are located on the H2AFZ gene, the amplified products can be allele detected using direct sequencing or other effective means. And selecting and reserving according to the SNP locus gene frequency result, and reserving seeds for chickens with high dominant SNP frequency.
The genotype of the polymorphic site contained in the marker rs737893701 is CC or TC or TT, and/or the genotype of the polymorphic site contained in the marker rs735176537 is AG or AA or GG, and/or the genotype of the polymorphic site contained in the marker rs736904000 is AG or AA or GG, and/or the genotype of the polymorphic site contained in the marker rs738535625 is AG or AA or GG, and/or the genotype of the polymorphic site contained in the marker rs80690045 is AG or AA or GG, and/or the genotype of the polymorphic site contained in the marker rs80765089 is TC or TT or CC.
Reserving seeds according to the number of the hens not less than 80 and the proportion of the hens to the hens not less than 1:10, and constructing a new family seed reproduction in the egg laying peak period. Screening according to dominant SNP frequency may improve disease resistance of chicken flock.
The foregoing description of some exemplary embodiments has been presented for purposes of illustration. Although the foregoing discussion has set forth specific embodiments, workers skilled in the art will recognize that changes may be made in form and detail without departing from the broader spirit and scope of the invention. The specification and drawings are, accordingly, to be regarded in an illustrative rather than a restrictive sense. The detailed description is, therefore, not to be taken in a limiting sense, and the scope of the present invention is defined only by the appended claims, along with the full scope of equivalents to which such claims are entitled.
Claims (4)
1. A method for screening SNPs on an integrated disease resistance trait candidate gene, the method comprising the steps of:
(1) Constructing a reference population and determining serum immune factor levels of both populations;
(2) Each chicken is sampled and preserved, DNA is extracted, and the SNP chip of Beijing core number one is utilized to carry out parting comprising the above 5 SNPs sites, and the data after genotyping is processed and the quality is controlled;
(3) Extracting peripheral blood macrophages of a 28-day-old uninfected Wenchang chicken (n=8) and a big-bone chicken (n=8) for transcriptome sequencing analysis;
(4) Transcriptome combined selection signal clearing analysis screens SNP loci which are obviously differentiated on candidate genes related to disease resistance.
2. A group of remarkable SNP markers related to disease resistance of broiler chickens and positioned on genes affecting disease resistance traits of chicken chromosome 4, which is characterized in that the SNP markers specifically comprise:
3. use of the SNP marker of claim 2, characterized in that the use is a method for selecting a clone with higher disease resistance comprising: obtaining genomic DNA from chicken and SNP chip-based whole genome SNPs; peripheral blood macrophage mRNA expression, screening candidate genes related to disease resistance by utilizing transcriptome expression difference and selective signal elimination analysis, screening out SNP (single nucleotide polymorphism) which is obviously differentiated on the candidate genes, and selecting individuals with high SNP gene frequency for seed reservation.
4. The use according to claim 3, wherein the selection of individuals with high frequency of SNP genes is: the genotype of the polymorphic site contained in the marker rs737893701 is CC or TC or TT, and/or the genotype of the polymorphic site contained in the marker rs735176537 is AG or AA or GG, and/or the genotype of the polymorphic site contained in the marker rs736904000 is AG or AA or GG, and/or the genotype of the polymorphic site contained in the marker rs738535625 is AG or AA or GG, and/or the genotype of the polymorphic site contained in the marker rs80690045 is AG or AA or GG, and/or the genotype of the polymorphic site contained in the marker rs80765089 is TC or TT or CC.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310607006.4A CN116606938A (en) | 2023-05-26 | 2023-05-26 | SNP locus related to improving innate immunity of yellow-feather broilers and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202310607006.4A CN116606938A (en) | 2023-05-26 | 2023-05-26 | SNP locus related to improving innate immunity of yellow-feather broilers and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN116606938A true CN116606938A (en) | 2023-08-18 |
Family
ID=87677815
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310607006.4A Pending CN116606938A (en) | 2023-05-26 | 2023-05-26 | SNP locus related to improving innate immunity of yellow-feather broilers and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116606938A (en) |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104404037A (en) * | 2014-11-17 | 2015-03-11 | 中国农业科学院北京畜牧兽医研究所 | Cultivation method of chicken disease-resistant new line and SNP (Single Nucleotide Polymorphism) marker related to chicken disease resistance |
| CN111225986A (en) * | 2017-10-10 | 2020-06-02 | 中国农业科学院北京畜牧兽医研究所 | Chicken whole genome SNP chip and application thereof |
| CN112002371A (en) * | 2020-07-31 | 2020-11-27 | 中国农业科学院北京畜牧兽医研究所 | Genome selection method for residual feed intake of white feather broilers |
| CN113549699A (en) * | 2021-07-01 | 2021-10-26 | 中国农业科学院北京畜牧兽医研究所 | Genome selection method for egg number of white feather broilers |
-
2023
- 2023-05-26 CN CN202310607006.4A patent/CN116606938A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104404037A (en) * | 2014-11-17 | 2015-03-11 | 中国农业科学院北京畜牧兽医研究所 | Cultivation method of chicken disease-resistant new line and SNP (Single Nucleotide Polymorphism) marker related to chicken disease resistance |
| CN111225986A (en) * | 2017-10-10 | 2020-06-02 | 中国农业科学院北京畜牧兽医研究所 | Chicken whole genome SNP chip and application thereof |
| CN112002371A (en) * | 2020-07-31 | 2020-11-27 | 中国农业科学院北京畜牧兽医研究所 | Genome selection method for residual feed intake of white feather broilers |
| CN113549699A (en) * | 2021-07-01 | 2021-10-26 | 中国农业科学院北京畜牧兽医研究所 | Genome selection method for egg number of white feather broilers |
Non-Patent Citations (3)
| Title |
|---|
| 刘冉冉;赵桂苹;文杰;: "鸡基因组育种和保种用SNP芯片研发及应用", 中国家禽, no. 15 * |
| 刘向萍;郭晓敏;李志腾;朱静;盛中伟;马腾;常国斌;窦新红;王克华;陈国宏;: "狼山鸡NLRC5基因启动子区SNP及其在先天免疫中的效应分析", 畜牧兽医学报, no. 01 * |
| 王统苗;郑圣晗;张莘;陈博雯;刘本帅;张扬;王志秀;常国斌;陈国宏;: "不同鸡品种BTN2基因SNP和INDEL分析", 四川农业大学学报, no. 03 * |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN106434931B (en) | A kind of structure variation SV177 and its detection technique distinguishing Large White and place of china pig variety | |
| CN106399523B (en) | A kind of molecular labeling SV193 and its application technology distinguishing Chinese native pig breed and Large White | |
| CN106701931B (en) | SNP marker related to rapid growth of Micropterus salmoides 'Youbei No. 1' and application thereof | |
| CN104673902B (en) | SNP molecular marker related to breast muscle weight and breast muscle percentage of chicken and application of SNP molecular marker | |
| CN104962634A (en) | Kit used for detecting body size traits of chicken and molecular breeding method of chicken | |
| CN109321665B (en) | Method for screening molecular markers of Jinhu black-bone chicken and application thereof | |
| CN106011259B (en) | Duolang sheep SNP marker and screening method and application thereof | |
| CN112609009B (en) | SNP molecular marker combination related to weight and body size of Guangxi three-yellow chicken based on whole genome sequencing screening and application | |
| CN108251418B (en) | SNP molecular marker related to growth speed of broiler chicken and application thereof | |
| CN111057771B (en) | SNP molecular marker for distinguishing 'Zhongyang No. 1' from common fugu obscurus and application thereof | |
| CN104894289A (en) | Kit for detecting chicken growth characters and chicken molecular breeding method | |
| CN116179714B (en) | Molecular marker related to chicken slaughtering and meat quality characteristics and breeding method of high-quality slaughtering and processing type novel variety | |
| CN108570508B (en) | Molecular marker related to chicken beak malformation character and application thereof | |
| CN116606938A (en) | SNP locus related to improving innate immunity of yellow-feather broilers and application thereof | |
| CN115261486A (en) | Huaxi cattle whole genome selective breeding chip and application thereof | |
| CN103710427A (en) | Single nucleotide polymorphism, detection method and application of chicken gene | |
| CN116103413B (en) | SNP (Single nucleotide polymorphism) marker related to laying characteristics of local chickens as well as detection method and application thereof | |
| CN108866210B (en) | Molecular marker related to red blood cell number of 20-day-old piglet and application thereof | |
| CN116751868B (en) | SNP (Single nucleotide polymorphism) marker related to laying characteristics of local chickens as well as detection method and application thereof | |
| CN117327809B (en) | Bay scallop accumulated carotenoid related molecular marker and application thereof | |
| CN116814798B (en) | Molecular marker for breeding rapid growth type largemouth black bass and application thereof | |
| CN115851962B (en) | Molecular marker related to abdominal fat weight character of chicken and application thereof | |
| CN111154849B (en) | Method for identifying size and ploidy of Acipenser dabryanus genome | |
| KR20130050832A (en) | Methods of genetic characteristics and cumulative power of discrimination in korean native chicken and korean native commercial chicken | |
| CN106498073A (en) | A kind of SNP marker related to Xinjiang Donkey milk production trait and its 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 |