CN115820875A - Molecular marker for evaluating pig eye muscle area and screening method and application thereof - Google Patents
Molecular marker for evaluating pig eye muscle area and screening method and application thereof Download PDFInfo
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Abstract
The invention relates to a molecular marker for evaluating the area of pig eye muscles, a screening method and application thereof, and relates to the technical field of pig genetic genes. The molecular marker is an SNP molecular marker, and is positioned at 30786798bp position of pig No. 7 chromosome, 120345012bp position of pig No. 7 chromosome, 56213488bp position of pig No. 14 chromosome, and/or 74418117bp position of pig No. 5 chromosome. The molecular marker is used for evaluating the pig eye muscle area and further evaluating the pig lean meat percentage, is used for breeding pigs, can effectively improve the pig eye muscle area and the pig lean meat percentage, further improves the breeding molecular marker of the pig ketone body shape, and provides an effective means for improving the pork quality and the lean meat yield.
Description
Technical Field
The invention relates to the technical field of pig genetic genes, in particular to a molecular marker for evaluating pig eye muscle area and a screening method and application thereof.
Background
China is a big pig-raising country and a big pork consumption country, pork is one of main animal protein sources of China all the time, and the pork yield is closely related to the stability and level of the life of people in China. The growth traits of lean meat percentage, daily gain, daily feed intake and the like are directly related to the protein production performance of the pork alone, the eye muscle area has high correlation with the lean meat percentage and the determination of the lean meat percentage is more convenient, so the eye muscle area becomes an important economic trait in the current breeding of the pork industry.
The eye muscle area and the lean meat percentage are in medium-high positive correlation, and the calculation of the lean meat percentage requires that the ratio of the lean meat to the carcass weight of the pig is calculated after the pig is slaughtered, so that the measurement of the lean meat percentage is complex and the cost is high, and the method is not suitable for being used as a selective retention index of conventional breeding. However, the area of the eye muscle can be measured by B-ultrasonic measurement between the third last and the fourth ribs of the pig at a position 5cm away from the dorsal midline. Therefore, the eye muscle area can be used as a selection index for replacing lean meat percentage to a considerable extent.
The eye muscle area is a quantitative trait controlled by multiple genes, and the like. For such quantitative traits, multiple detection of candidate genes and QTLs associated with the trait contributes to the accuracy of genome selection. Most QTLs are identified using linkage disequilibrium, and the identified QTLs are difficult to use for genetic improvement in swine.
Disclosure of Invention
Aiming at the problems, the invention provides a molecular marker for evaluating the pig eye muscle area, and the evaluation of the pig eye muscle area is carried out according to the molecular marker, so that the pig lean meat percentage is evaluated, and the molecular marker is used for breeding pigs, can effectively improve the pig eye muscle area, improve the pig lean meat percentage, further improve the breeding molecular marker of the pig ketone body shape, and provides an effective means for improving the pork quality and the lean meat yield.
In order to achieve the purpose, the invention provides a molecular marker for evaluating the pig eye muscle area, which is an SNP molecular marker, and the molecular marker is located at 30786798bp position of pig chromosome 7, 120345012bp position of pig chromosome 7, 56213488bp position of pig chromosome 14 and/or 74418117bp position of pig chromosome 5.
In one embodiment, the molecular marker is located at 30786798bp of pig chromosome 7, and when the genotype of the molecular marker is CC, the area of the eye muscle of the pig to be evaluated is high;
the molecular marker is located at 120345012bp of the pig No. 7 chromosome, and when the genotype of the molecular marker is CC, the high eye muscle area of the pig to be evaluated is prompted;
the molecular marker is located at 56213488bp of pig No. 14 chromosome, and when the genotype of the molecular marker is CC, the high eye muscle area of the pig to be evaluated is prompted;
and/or the molecular marker is positioned at 74418117bp of the pig No. 5 chromosome, and when the genotype of the molecular marker is TT, the area of the eye muscle of the pig to be evaluated is high.
The different genotypes of the molecular marker have obvious difference in the area of the eye muscles of pigs, wherein when the molecular marker is positioned at the 30786798bp position of the chromosome 7 of the pig, the area of the eye muscles between individuals of the genotype CC and the TC and TT Duroc boar is obviously improved by 1.39 percent and 1.97 percent, so that C in the marker is an allele which is beneficial to obviously improving the area of the eye muscles; when the molecular marker is located at the 120345012bp position of the pig No. 7 chromosome, the genotype CC is obviously improved by 3.18 percent compared with the TT, so the C in the marker is an allele which is beneficial to improving the eye muscle area; when the molecular marker is positioned at the 56213488bp position of the No. 14 pig chromosome, the genotype CC is obviously improved by 2.46 percent compared with the TT duroc boar individual interocular muscle area, so that C is more beneficial to T and improves the ocular volume area in the marker; when the molecular marker is located at 74418117bp of pig chromosome 5, the gene type TT is respectively improved by 1.59 percent and 3.07 percent compared with the eye muscle area between CT and CC duroc boar individuals, so that T in the gene is more beneficial to the improvement of the eye muscle area compared with C.
The invention also provides a screening method of the molecular marker, which comprises the following steps:
collecting phenotype data: acquiring phenotype data of the eye muscle area of a pig to be detected;
obtaining SNP molecular markers: sampling a pig to be detected, extracting DNA, and genotyping to obtain an SNP molecular marker covering the whole genome; carrying out physical position updating and quality control on the SNP molecular marker covering the whole genome;
screening SNP molecular markers: and (3) carrying out GWAS analysis on the SNP molecular marker after quality control by combining phenotype data, analyzing the difference condition of the eye muscle areas of pigs with different genotype groups, and screening to obtain the SNP molecular marker for evaluating the eye muscle areas of the pigs.
With the development of sequencing technology in recent years, a large amount of sequencing information can be obtained, genome-wide association analysis (GWAS) is widely applied to animal breeding research, the development of chip sequencing technology enables the acquisition of high-density SNP information to be more convenient, the related research of GWAS is more convenient, and the GWAS can effectively identify SNP which is obviously related to traits. The research uses a farmCPU model in rMVP software for analysis, and the model alternately uses a fixed effect and a random effect, can quickly and accurately identify the obvious SNP marker, and has promotion significance on genetic improvement of the eye muscle area.
The invention also provides a detection kit for evaluating the lean meat percentage of the pigs, which comprises a reagent for detecting the molecular marker.
The invention also provides a method for evaluating the lean meat percentage of the pig, which comprises the steps of obtaining the detection result of the molecular marker, and judging the eye muscle area of the pig according to the detection result so as to judge the lean meat percentage of the pig.
In one embodiment, the molecular marker is located at 30786798bp of pig chromosome 7, and when the genotype of the molecular marker is CC, the area of the eye muscle of the pig is judged to be high, and the lean meat percentage of the pig is judged to be high;
the molecular marker is located at 120345012bp of the No. 7 chromosome of the pig, and when the genotype of the molecular marker is CC, the eye muscle area of the pig is judged to be high, and the lean meat percentage of the pig is judged to be high.
In one embodiment, the molecular marker is located at 56213488bp of pig chromosome 14, and when the genotype of the molecular marker is CC, the area of the eye muscle of the pig is judged to be large, and the lean meat percentage of the pig is judged to be high.
In one embodiment, the molecular marker is located at 74418117bp of pig chromosome 5, and when the genotype of the molecular marker is TT, the area of the eye muscle of the pig is judged to be high, and the lean meat percentage of the pig is judged to be high.
The invention also provides a breeding method of pigs, detection is carried out according to the method, the lean meat percentage of the pigs is judged according to the genotype of the molecular marker, and the pigs with high lean meat percentage are reserved; the molecular marker is located at 30786798bp position of pig chromosome 7, 120345012bp position of pig chromosome 7, 56213488bp position of pig chromosome 14 or 74418117bp position of pig chromosome 5.
In one embodiment, the molecular marker is located at 30786798bp of pig chromosome 7, and when the genotype of the molecular marker is CC, the area of eye muscles of a pig is judged to be high, the lean meat percentage of the pig is judged to be high, and the pig is reserved;
the molecular marker is located at 120345012bp of the No. 7 pig chromosome, and when the genotype of the molecular marker is CC, the eye muscle area of the pig is judged to be high, the lean meat percentage of the pig is judged to be high, and the pig is reserved;
the molecular marker is located at 56213488bp of No. 14 chromosome of the pig, and when the genotype of the molecular marker is CC, the eye muscle area of the pig is judged to be high, the lean meat percentage of the pig is judged to be high, and the pig is reserved;
the molecular marker is located at 74418117bp of pig chromosome 5, and when the genotype of the molecular marker is TT, the eye muscle area of the pig is judged to be high, the lean meat percentage of the pig is judged to be high, and the pig is reserved.
Compared with the prior art, the invention has the following beneficial effects:
according to the molecular marker for evaluating the pig eye muscle area, the pig lean meat percentage is evaluated according to the molecular marker, the molecular marker is used for breeding pigs, the pig eye muscle area can be effectively increased, the pig lean meat percentage is increased, the breeding molecular marker of the pig ketone body shape is further perfected, and an effective means is provided for improving the pork quality and the lean meat yield.
Drawings
FIG. 1 is a Manhattan diagram of marker screening in the examples.
Detailed Description
To facilitate an understanding of the invention, the invention will now be described more fully with reference to the accompanying drawings. Preferred embodiments of the present invention are shown in the drawings. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. The terminology used in the description of the invention herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. As used herein, the term "and/or" includes any and all combinations of one or more of the associated listed items.
Defining:
SNP molecular markers: refers to the variation of a single nucleotide on the genome, resulting in a genetic marker.
The source is as follows:
GGP 50k SNP(GeneSeek，US)。
reagents, materials and equipment used in the embodiment are all commercially available sources unless otherwise specified; unless otherwise specified, all the experimental methods are routine in the art.
Examples
1. Phenotypic data is collected.
The research group of the invention is 2793 Duroc boars, all pigs are from a breeding farm of Yangxi Yangxiang GmbH company in 2014-2021, the measuring part of the eye muscle area is 5cm away from the back midline between the third to the fourth ribs of the penultimate pig, and the Mylab Touch Vet B ultrasonic instrument is used for measuring the weight of the pigs at the stage of 100-135 kg. The measured data were corrected to 115kg body weight levels by the KFNets system, and 2783 duroc boars were left for association analysis after the corrected data were quality controlled by the mean ± 3-fold standard deviation.
2. And acquiring the SNP molecular marker.
Collecting ear tissue sample or blood sample of pig, extracting total DNA, genotyping by GGP 50k SNP chip, and shapingAnd synthesizing multiple sequencing results to obtain 50697 SNP markers covering the whole genome. The physical location of all SNP markers was updated according to the latest version of the pig reference genome (Sscofa 11.1) using the NCBI genome alignment program (https:// www.ncbi.nlm.nih.gov /). SNPs with unknown genomic positions and SNPs on sex chromosomes are not used for association analysis. For SNP markers on all autosomes, quality control was performed using the Plink software, with the criteria: the individual detection rate is more than or equal to 90 percent; the SNP detection rate is more than or equal to 90 percent; the minimum allele frequency is more than or equal to 0.01; the Hardy-Weinberg equilibrium p value is more than or equal to 10 -6 . Filling individuals with the deleted genotypes by adopting Beagle software (version 4.1), and controlling the quality again after filling, wherein the quality control conditions are the same as the above conditions. And after quality control, the remaining 39428 SNP sites are used for association analysis with the eye muscle area.
3. And (4) screening SNP molecular markers.
1. And (5) carrying out a statistical model.
Using a multi-label correlation model method, taking field-year-season and column as fixed effects, and using a FarmCPU model of an rMVP software package to perform GWAS analysis in an R statistical environment, wherein the research model is as follows:
Y n ＝T ni W i +P nj Q j +e n
wherein, Y n A table-type value vector, T, representing the nth individual ni The gene type of field-year-season, column and i pseudo QTNs and the first three main components for controlling the genetic background of the population are fixed effects; w i Representing the corresponding effect; p nj A jth tag representing an nth individual; q j Represents the jth corresponding effect; e.g. of the type n Denotes a residual vector, and follows a normal distribution, e to N (0,I σ 2 e), and σ 2e denotes a residual variance. P value is 1 divided by all SNP site numbers, i.e.: p =1/39428.
2. And (4) marker screening.
And for the p values of all the markers, taking the negative logarithm conversion value of the markers to draw a Manhattan graph, and displaying and screening the most significant SNP markers. And analyzing the eye muscle area difference of different gene type group Duroc boars by using variance analysis (field-year-season, field and corresponding obvious SNP site effect are considered in the model) and Duncan multiple comparison (R statistical analysis platform) to analyze the eye muscle area difference of different gene type groups of the ASGA0032562, the ASGA0036920, the H3GA0040331 and the ALGA 0032929.
The results of the analysis are shown in FIG. 1 and the following table.
TABLE 1 ASGA0032562 labeling of different genotype pig eye muscle areas
TABLE 2 ASGA0036920 markers for different genotypes of porcine eye muscle area
TABLE 3 H3GA0040331 labeling different genotype porcine eye muscle areas
TABLE 4 ALGA0032929 labeling porcine eye muscle areas of different genotypes
The invention identifies a molecular marker for evaluating the pig eye muscle area: ASGA0032562, ASGA0036920, H3GA0040331 or ALGA0032929, which marked very significant differences in eye muscle area of pigs of different genotypes.
The ASGA0032562 marker is located at the 30786798bp position of the pig No. 7 chromosome, and the position is T > C mutation. The upstream and downstream 200bp gene sequences of the SNP marker are as follows, wherein E is the site of the SNP molecular marker:
5’-CTTGCCYTTTCTAGTGCACCTTTTCAGTATCTGCCAGAGTAACAGAGAAGGAAGAATT CCATCCTTCATTCAACAAATCATTTTTATTTATTTATTTGGCTGTSCCCACAGCATGTAAAAKTTCCAGCCAGGGATCAAACTCAAGCCATAGCAGTGACGCCAGATCTTTAACAGGGCCAGGGATYGAACCYGCATGGATG(SEQ ID NO:1)-E-TATTCGGGTCTACTAGCTACTGAGCC ATGACAGGAGCTCCTATTTTTTGCTTTTTAGGGCTATATCTGCAGCATATGGAAGTTCCCAGGCTAGGGGTCGAGTCAGAGCTGTAGCTGCCAGCCTACACCACAGCCACAGCAACACC AGATCAGAACTACATCTGTGACCTCCACCACTGCTCTCAGCAATGCTRGATCCTT-3’(SEQ ID NO:2)。
the ASGA0036920 marker is located at 120345012bp of pig chromosome 7, and the site is T > C mutation. The upstream and downstream 200bp gene sequences of the SNP marker are as follows, wherein F is the site of the SNP molecular marker:
5’-GTCCCCACAGCTGCACTGCCAGTCAGGGTTCTGCAGGCAGAGCYCRWAGGATGTATG CACACGGGTATATAAGGGGCTTTGTCCYAAGGAATTATTTCTCATGCCCATGGAGGTGGGCAAGGWGAGCTGATGTTTCCATTGRAATCAAAGGCAGGAGAAAGCCRATGTCCCAGCT CAAGGGCAGTCAGAMAGGGGGARCT(SEQ ID NO:3)-F-CCACTTACTCCAGGGAGGATGAGCCTTTGGTTTTACTCAGGCCCTCAACTGATTGGGTGAGGCCCACCCACCKGGGGACG GGCAATCTGCTTWYTTCTTAAAGGTTCATCTCACCCCATATATCCTCATAGGAMCCCCCC CCCCCSAGAATCATGCTGACCATATGTCTTGGACACCCCATGACCCAGCCAAACTGACAC -3’(SEQ ID NO:4)。
the H3GA0040331 marker is located at 56213488bp of pig chromosome 14, and the site is C > T mutation. The upstream and downstream 200bp gene sequences of the SNP marker are as follows, wherein L is the site of the SNP molecular marker:
5’-YGGAATCCAAGTCATGTCTTTGACCTTACACCACAGCTCATGGCAATGCTGGATCCTT GACYCACTGAGCGAGRCCAGGGATTAAACCTGCATCCTCATGGATACTAKTYGGGTTTGTTACTGCYGAGCCACAGTGGGAAMTCCTCAAGCCCACTTTTAAGTCCCACGTTTCTGCC CTCTCGTAGGCTGCCTCTCGAAGA(SEQ ID NO:5)-L-CAGGTGGGAGGATGTGGGCCTTACGGAGGCGACTGGGAGCATGGGAGGAACAGATGGCACAGGTCTCTCTCCAGTACAT ACCAGTCACTACCTGGAAGCTAGCAGCTATGATGTAACYACCTTGTCCGGTGAAGGCTG CCTTTGCTCTGTGGGTGGGACYGCGATGCCGCCAGGCTGGGGAATCCCATTGGTGACAA CAC-3’(SEQ ID NO:6)。
the ALGA0032929 marker is located at 74418117bp position of pig chromosome 5, which is the A > G mutation. The upstream and downstream 200bp gene sequences of the SNP marker are as follows, wherein O is the site of the SNP molecular marker:
5’-AGCTTGATTTTGTAATTTTTCTCCTTTATTAATKGCCTCTTTTTTTTTGGTAAGACTTGT ACTAAGAASTCATGATAAAAAGATGGTCCATGCCCTCGAGGAGCTTACATCTGATGTTACTCCTTGTCTTATTTTGATCCACCTAAATRAAATCCACACYGAAGGATTACAAACTCTAAG CCTAAAGCTGYGGTTTTTAA(SEQ ID NO:7)-O-TGGGAAAGATTTTGTGYCTAGGAGA TATTTGGTAAMACCTGGAGATATTTTTGGTTGTAACAATTGGAGGATGCTACTAGCATCTAGTGGGGAGAAACCAGARATGGTGCTAAACATCCTAAAATGCACAGGGCAGCTCTTCTC CCCTAAYSATRCAGAATTATCCAGCCTCAAAATCAAAACTTCTGAGGTTGAGAAAT-3’(SEQ ID NO:8)。
r, Y, M, K, S, W in the above sequence is a mixed base symbol, and the meaning is as follows:
r represents G or A, Y represents T/U or C, M represents A or C, K represents G or T/U, S represents G or C, and W represents A or T/U.
The ASGA0032562 marker genotype CC is obviously improved by 1.39 percent and 1.97 percent compared with the TC and TT duroc boar individuals, so C in the marker is an allelic group which is favorable for obviously improving the eye muscle area; the ASGA0036920 marker genotype CC is obviously improved by 3.18% compared with TT, so C in the marker is an allele which is beneficial to improving the eye muscle area; the H3GA0040331 marker genotype CC is obviously improved by 2.46 percent compared with the area of the interocular muscle of TT duroc boar individuals, so that C in the marker is more beneficial to T and improves the area of the ocular area; the ALGA0032929 marker genotype TT is improved by 1.59 percent and 3.07 percent respectively compared with the eye muscle area between CT and CC duroc boars, so that T in the gene is more beneficial to the improvement of the eye muscle area compared with C.
By detecting and marking ASGA0032562, ASGA0036920, H3GA0040331 and ALGA0032929 genotypes to assist breeding of breeding pigs, the Duroc boars with CC, CC and TT genotypes can be respectively selected and retained, the eye muscle area is increased, and the improvement of the eye area of the boars is facilitated, so that the lean meat percentage is increased.
The technical features of the embodiments described above may be arbitrarily combined, and for the sake of brevity, all possible combinations of the technical features in the embodiments described above are not described, but should be considered as being within the scope of the present specification as long as there is no contradiction between the combinations of the technical features.
The above-mentioned embodiments only express several embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. Therefore, the protection scope of the present patent shall be subject to the appended claims.
Claims (10)
1. A molecular marker for evaluating the pig eye muscle area is characterized in that the molecular marker is an SNP molecular marker, and the molecular marker is located at 30786798bp position of pig chromosome 7, 120345012bp position of pig chromosome 7, 56213488bp position of pig chromosome 14 and/or 74418117bp position of pig chromosome 5.
2. The molecular marker of claim 1, wherein the molecular marker is located at 30786798bp of pig chromosome 7, and when the genotype of the molecular marker is CC, the molecular marker indicates that the eye muscle area of a pig to be evaluated is high;
the molecular marker is located at 120345012bp of the pig No. 7 chromosome, and when the genotype of the molecular marker is CC, the high eye muscle area of the pig to be evaluated is prompted;
the molecular marker is located at 56213488bp of pig No. 14 chromosome, and when the genotype of the molecular marker is CC, the high eye muscle area of the pig to be evaluated is prompted;
and/or the molecular marker is positioned at 74418117bp of the pig No. 5 chromosome, and when the genotype of the molecular marker is TT, the area of the eye muscle of the pig to be evaluated is high.
3. A method for screening a molecular marker according to any one of claims 1 to 2, comprising the steps of:
collecting phenotype data: acquiring phenotype data of the eye muscle area of a pig to be detected;
obtaining SNP molecular markers: sampling a pig to be detected, extracting DNA, and genotyping to obtain an SNP molecular marker covering the whole genome; carrying out physical position updating and quality control on the SNP molecular marker covering the whole genome;
screening SNP molecular markers: and (3) carrying out GWAS analysis on the SNP molecular marker after quality control by combining phenotype data, analyzing the difference condition of the eye muscle areas of pigs with different genotype groups, and screening to obtain the SNP molecular marker for evaluating the eye muscle areas of the pigs.
4. An assay kit for assessing lean meat percentage in a pig comprising reagents for detecting the molecular marker of any one of claims 1-2.
5. A method for evaluating lean meat percentage of swine, comprising obtaining a result of detecting the molecular marker of any one of claims 1-2, and determining an eye muscle area of the swine based on the result of the detection, thereby determining the lean meat percentage of the swine.
6. The method of claim 5, wherein the molecular marker is located at 30786798bp of pig chromosome 7, and when the genotype of the molecular marker is CC, the eye muscle area of the pig is judged to be high, and the pig lean meat percentage is judged to be high;
the molecular marker is located at 120345012bp of the pig No. 7 chromosome, and when the genotype of the molecular marker is CC, the eye muscle area of the pig is judged to be large, and the lean meat percentage of the pig is judged to be high.
7. The method of claim 5, wherein the molecular marker is located at 56213488bp of chromosome 14 of pig, and when the genotype of the molecular marker is CC, the eye muscle area of pig is judged to be large, and the pig lean meat percentage is judged to be high.
8. The method of claim 5, wherein the molecular marker is located at 74418117bp of pig chromosome 5, and when the genotype of the molecular marker is TT, the eye muscle area of the pig is judged to be high, and the pig lean meat percentage is judged to be high.
9. A breeding method of pigs, characterized in that the method according to any one of claims 5-8 is used for detection, the lean meat percentage of the pigs is judged according to the genotype of the molecular marker, and the pigs with high lean meat percentage are reserved; the molecular marker is located at 30786798bp position of pig chromosome 7, 120345012bp position of pig chromosome 7, 56213488bp position of pig chromosome 14 or 74418117bp position of pig chromosome 5.
10. The breeding method according to claim 9, wherein the molecular marker is located at 30786798bp of pig chromosome 7, and when the genotype of the molecular marker is CC, the eye muscle area of the pig is judged to be high, the lean meat percentage of the pig is judged to be high, and the pig is reserved;
the molecular marker is located at 120345012bp of the No. 7 pig chromosome, and when the genotype of the molecular marker is CC, the eye muscle area of the pig is judged to be high, the lean meat percentage of the pig is judged to be high, and the pig is reserved;
the molecular marker is located at 56213488bp of a 14-th chromosome of the pig, and when the genotype of the molecular marker is CC, the eye muscle area of the pig is judged to be high, the lean meat percentage of the pig is judged to be high, and the pig is reserved;
the molecular marker is located at 74418117bp of pig chromosome 5, and when the genotype of the molecular marker is TT, the eye muscle area of the pig is judged to be high, the lean meat percentage of the pig is judged to be high, and the pig is reserved.
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