CN116515956A - Molecular marker for screening goose eggs based on GWAS and selection method - Google Patents
Molecular marker for screening goose eggs based on GWAS and selection method Download PDFInfo
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Abstract
The invention discloses a method for screening goose egg molecular markers and selecting based on GWAS, which relates to the field of gene detection, and the technical scheme of the invention comprises the following steps: genome sequencing and time-of-flight mass spectrometry detection are carried out on a plurality of female goose measuring groups in the generation 5; and carrying out time-of-flight mass spectrometry genotype detection on a plurality of breeding groups of 7-generation geese. The egg laying number of the 7 generation can be obviously improved, and the genetic progress is obviously higher than that of the first two generations.
Description
Technical Field
The invention relates to a molecular marker for screening goose eggs based on GWAS and a selection method, and mainly relates to the field of gene detection.
Background
Is a seasonal breeding poultry, and compared with chickens and ducks, the method has the advantages of late egg laying and less egg laying, so that the selection of the egg laying amount in breeding is of great practical significance. Egg laying performance belongs to quantitative traits, is controlled by micro-effect multiple genes, and the traditional method has low accuracy and needs to improve genetic progress by using molecular marker assisted selection and other technologies.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides a molecular marker and a selection method for screening goose eggs based on GWAS, which can obviously improve the number of eggs laid in 7 generations and lead the genetic progress to be obviously higher than that of the first two generations.
In order to achieve the above purpose, the technical scheme of the invention is as follows:
a method for screening goose egg-laying molecular markers based on GWAS comprises the following steps:
genome sequencing and time-of-flight mass spectrometry detection are carried out on a plurality of female goose measuring groups in the generation 5;
and carrying out time-of-flight mass spectrometry genotype detection on a plurality of breeding groups of 7-generation geese.
Preferably, 2mL of blood from the pteran vein is collected by a vacuum blood collection tube containing heparin sodium, and the blood is preserved at-20 ℃ for later use, wherein the egg yield measurement time is from the start of egg yield to 60/66 weeks of age.
Preferably, the blood genomic DNA is extracted using a blood genomic extraction kit, and after passing the quality control by a NanoDrop2000 spectrophotometer, the whole gene re-sequencing is performed using the IlluminaHiSeqXTen platform.
Preferably, after data filtration, the BWA software compares to the goose chromosome genome and the GATK software performs SNP genotype data detection; the Plink software performs quality control on the obtained SNP data, detects the layering condition of the group structure by a principal component analysis method, and performs GWAS analysis by using GEMMA software.
Preferably, whole genome correlation analysis is performed using a mixed linear model of GEMMA software y=wα+xβ+epsilon, wald tests for significance between SNPs markers and multiplex validation using Bonferroni's method;
y is the phenotype value of the individual, W is the population structure, x is the genotype of the SNP, beta is the SNP marker effect value, epsilon random residuals.
Preferably, the significantly associated SNPs identified by GWAS are validated using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The general linear model of JMP software, y=μ+x+epsilon, calculates the least squares average for each genotype and analyzes the differences between genotypes, which were checked with Bonferroni;
y is the phenotypic value, average μ population, x genotype, epsilon random error.
Selecting a goose egg-laying molecule selection method based on GWAS, primarily selecting 600 female geese and 150 male geese according to the egg yield of parent families, and detecting haplotypes and haplotype combinations constructed by the 4 SNP by adopting a time-of-flight mass spectrometry method; screening dominant haplotype combined male goose individuals and female goose individuals, and combining phenotypes to select 340 female geese, 86 male geese and other female goose individuals containing the dominant haplotype to form a measurement group together for cage loading measurement.
The technical principle and the beneficial effects of the invention are as follows:
after the determination and selection by the method, the egg laying number of 7 generations can be obviously improved, and the genetic progress is obviously higher than that of the first two generations.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings required for the description of the embodiments will be briefly described, and it is obvious that the drawings in the following description are only two of the present invention, and other drawings can be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 shows that the 4 SNPs associated with 66 week-old egg production are in 1 haplotype block;
FIG. 2 shows the results of time-of-flight mass spectrometry detection of 4 SNP loci;
FIG. 3 shows the statistics of haplotype numbers and haplotype frequencies;
FIG. 4 shows the distribution of the haplotype combinations and the statistics of egg yield;
FIG. 5 shows the distribution results of the haplotype combination detection of 7-generation parent geese;
FIG. 6 shows the egg yield of each haplotype combination female goose in 7 generations.
Detailed Description
The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings, and it is obvious that the described embodiments are only preferred embodiments of the present invention, not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Examples
A method for screening goose egg laying molecular markers based on GWAS comprises the following steps:
experimental animal and egg production trait determination
Taking 209 parent geese of 5-generation part of Yuzhou white goose I line as a research object, carrying out genome re-sequencing and time-of-flight mass spectrometry detection, and carrying out time-of-flight mass spectrometry genotype detection by taking 750 geese of 7-generation breeding group as a research object. 2mL of blood of the pterygoid vein is collected by using a vacuum blood collection tube containing heparin sodium and is preserved at the temperature of minus 20 ℃ for standby. The egg production is measured by feeding individual cages (600 mm multiplied by 800mm multiplied by 900 mm), counting the egg production number every day from the start of production to 60/66 weeks of age, and freely drinking water and feeding standard feed.
GWAS association and haplotype analysis
Blood genome DNA was extracted using a blood genome extraction kit (Beijing Tiangen Co., DP 332), and after passing the quality control by a NanoDrop2000 spectrophotometer, the whole gene was re-sequenced using a Illumina HiSeq X Ten platform by the Tianjin Northa Biol Co., ltd. After data filtration, BWA software compares to a goose chromosome genome, and GATK software detects SNP genotype data; the Plink software performs quality control (parameter setting: geno 0.1;mind 0.1;MAF 0.05;hwe0.0000001) and principal component analysis on the obtained SNP data to detect population structure layering, and performs GWAS analysis by using GEMMA software.
Full genome association analysis was performed using a mixed linear model of GEMMA software, y=wα+xβ+epsilon (y is the phenotype value of the individual, W is the population structure, x is the genotype of the SNP, β is the SNP marker effect value, epsilon random residuals), wald examined the significance between SNPs markers and multiple-validated using Bonferroni's method (threshold: 10-7). The significantly associated SNPs identified by GWAS were verified using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), with a sequencing platform of MassARRAY genotyping. GAP software package draws whole genome association analysis Manhattan diagram under R language environment. The general linear model of JMP (13.0) software y=μ+x+epsilon (y is the phenotype value, μ population average, x genotype, epsilon random error) calculates the least squares average for each genotype and analyzes the differences between genotypes, which were checked with Bonferroni.
GWAS analysis and screening of molecular markers related to egg production number
Genome-wide resequencing yielded 2.896Tb data altogether, with an average depth of coverage of sequencing of 12.44X. After data filtration, 2.891Tb high-quality sequencing data map is compared with a goose reference genome sequence, the comparison rate is 96.58-98.38%, and 9,279,339 SNPs are detected in total. As a result of the GWAS study, it was found that 4 SNPs (SNP 17, SNP18, SNP281 and SNP 282) sites in chromosome 35 and haplotypes constructed by the same were potential molecular markers of 48 and 60 week egg numbers (FIG. 1). The 4 SNPs and the haplotype modules constructed by the SNPs are obviously related to the egg laying property and are positioned in a coding region of a transmembrane protein 161A (TMEM 161A) gene.
Different genotype detection results and egg yield statistical analysis
In the 5 th generation of the population, the genotype frequencies of the SNP loci and haplotypes were detected by the method of time-of-flight mass spectrometry (FIG. 2). The results showed that 11 haplotypes were detected in total (fig. 3), with CAGT (69.86%), CGGT (7.66%), TGAA (6.22%) and TAGT (5.74%) being dominant haplotypes; the 11 haplotypes together consisted of 19 haplotype combinations (FIG. 4).
Based on the research results, the 7-generation breeding process is performed by adopting a mode of combining molecular marker assisted selection with traditional breeding. According to the egg yield of the parent family, 600 female geese and 150 male geese are initially selected, haplotypes constructed by the 4 SNP and haplotype combination chart 5 are detected by adopting a time-of-flight mass spectrometry method, dominant haplotype combination male geese and female geese individuals are screened, 340 female geese, 86 male geese and other female geese individuals containing dominant haplotypes are selected from the obtained single-fold combinations, and the combined phenotypes form a measurement group together for cage loading measurement. The results of the egg production number measurement of each genotype are shown in FIG. 6, and the results of the measurement show that after selection, the egg production number of the 7 th generation at the age of 66 weeks is improved by 1.5 times compared with that of the 6 th generation, and the genetic progress is obviously higher than that of the first two generations (the 6 th generation is improved by 0.6 times and the 5 th generation is improved by 0.7 times).
According to the scheme, whole genome re-sequencing is carried out on a specific goose group, a Sichuan white goose high-quality whole genome SNP variation dataset is constructed, phenotype data of accurate egg production traits are acquired by combining intelligent measurement and data acquisition equipment, whole genome association analysis of egg production numbers is carried out, molecular markers of the egg production numbers of geese are screened, and molecular marker auxiliary selection is further carried out by screening dominant haplotypes of the egg production numbers.
The foregoing description of the preferred embodiments of the invention is not intended to be limiting, but rather is intended to cover all modifications, equivalents, alternatives, and improvements that fall within the spirit and scope of the invention.
Claims (7)
1. The method for screening the goose egg laying molecular marker based on the GWAS is characterized by comprising the following steps of:
genome sequencing and time-of-flight mass spectrometry detection are carried out on a plurality of female goose measuring groups in the generation 5;
and carrying out time-of-flight mass spectrometry genotype detection on a plurality of breeding groups of 7-generation geese.
2. The GWAS-based goose egg production molecular marker screening method according to claim 1, wherein the method is characterized in that: 2mL of blood of the pterygoid vein is collected by using a vacuum blood collection tube containing heparin sodium and is preserved at the temperature of minus 20 ℃ for standby, wherein the egg yield measurement time is from the start of egg yield to 60/66 weeks of age.
3. The GWAS-based goose egg production molecular marker screening method according to claim 1, wherein the method is characterized in that: and extracting blood genome DNA by using a blood genome extraction kit, and performing total gene re-sequencing by using an IlluminaHiSeqXTen platform after the quality inspection of the NanoDrop2000 spectrophotometer.
4. The GWAS-based goose egg production molecular marker screening method according to claim 1, wherein the method is characterized in that: after data filtration, BWA software compares to a goose chromosome genome, and GATK software detects SNP genotype data; the Plink software performs quality control on the obtained SNP data, detects the layering condition of the group structure by a principal component analysis method, and performs GWAS analysis by using GEMMA software.
5. The GWAS-based goose egg production molecular marker screening method according to claim 1, wherein the method is characterized in that: performing genome-wide association analysis using a hybrid linear model y=wα+xβ+ε of GEMMA software, wald testing for significance between SNPs markers and multiplex-testing using Bonferroni's method;
y is the phenotype value of the individual, W is the population structure, x is the genotype of the SNP, beta is the SNP marker effect value, epsilon random residuals.
6. The GWAS-based goose egg production molecular marker screening method according to claim 1, wherein the method is characterized in that: significant associations of SNPs identified by GWAS were verified using matrix-assisted laser desorption ionization time-of-flight mass spectrometry. The general linear model of JMP software, y=μ+x+epsilon, calculates the least squares average for each genotype and analyzes the differences between genotypes, which were checked with Bonferroni; y is the phenotypic value, average μ population, x genotype, epsilon random error.
7. The GWAS-based goose egg production molecular marker selection method according to claim 1, wherein the method comprises the following steps: 600 female geese and 150 male geese are initially selected according to the egg yield of the parent family, and haplotypes and haplotype combinations constructed by the 4 SNP are detected by adopting a time-of-flight mass spectrometry method; screening dominant haplotype combined male goose individuals and female goose individuals, and combining phenotypes to select 340 female geese, 86 male geese and other female goose individuals containing the dominant haplotype to form a measurement group together for cage loading measurement.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112592983A (en) * | 2020-12-23 | 2021-04-02 | 重庆市畜牧科学院 | SNP molecular marker method and primer for screening goose egg quality |
| US20210395803A1 (en) * | 2018-09-29 | 2021-12-23 | China Agricultural University | Egg-type chicken whole-genome snp chip and use thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| US20210395803A1 (en) * | 2018-09-29 | 2021-12-23 | China Agricultural University | Egg-type chicken whole-genome snp chip and use thereof |
| CN112592983A (en) * | 2020-12-23 | 2021-04-02 | 重庆市畜牧科学院 | SNP molecular marker method and primer for screening goose egg quality |
Non-Patent Citations (2)
| Title |
|---|
| GUANGLIANG GAO 等: "Genome-Wide Association Study-Based Identification of SNPs and Haplotypes Associated With Goose Reproductive Performance and Egg Quality", FRONT GENET, vol. 12 * |
| 胡彦竞科;张克山;钟航;王启贵;刘安芳;: "鹅GnIH基因多态性及其与产蛋量的关联性研究", 中国畜牧杂志, no. 21, pages 17 - 19 * |
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