CN115896299A - PSMD3 gene molecular marker related to chicken skin color character and carcass character and application thereof - Google Patents
PSMD3 gene molecular marker related to chicken skin color character and carcass character and application thereof Download PDFInfo
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Abstract
The invention discloses a PSMD3 gene molecular marker related to chicken skin color traits and carcass traits and application thereof, belonging to the technical field of biology. The molecular marker is positioned on a chicken PSMD3 gene, and the accession number of the PSMD3 gene is GeneID 426133; the molecular marker is specifically shown as SEQ ID NO:1, wherein the sequence exists NC-052558.1: G4304798A > G, NC _052558.1: G4304889G > A, NC _052558.1: g4304903C > T. Experiments prove that the 3 molecular markers are obviously related to the skin color character and carcass character of the chicken. Therefore, the method can accurately identify the chicken skin color character and the carcass character, and provides a new molecular marker for molecular marker-assisted selective breeding.
Description
Technical Field
The invention relates to the technical field of biology, in particular to a PSMD3 gene molecular marker related to chicken skin color traits and carcass traits and application thereof.
Background
With the improvement of living standard, the adjustment of policy and the development of industry, the demand of consumers for the iced fresh chicken is increased. Consumers prefer chicken with a yellow skin color, a bright skin color, and uniform color. The first impression that iced chicken present to consumers is skin color, and the shade and uniformity of skin color has become an important feature in determining consumer acceptance and broiler quality. The skin of poultry mostly appears yellow, and the color substances of the poultry are carotenoid and lutein. At present, most of the practical applications of chicken skin color research stay at the nutrition regulation level, the researches related to gene regulation and molecular markers are still in the continuous exploration and verification stage, and the breeding method is a breeding technical difficulty in how to breed the chickens with high yellowness, uniform color and stability, so that the skin color of the chickens meets the selection requirements of consumers. Single Nucleotide Polymorphism (SNP) is used as a DNA Molecular marker, has the characteristic of stable inheritance, can be used for Molecular marker-assisted Selection (MAS), has obvious effect on improving the breeding efficiency, and can be widely used for large-group large-scale breeding and screening of broiler chickens. Therefore, a molecular marker related to the skin yellowness of the chicken is searched, a breeding method for the skin yellowness of the chicken is developed, and the method has great significance for promoting the development of high-quality broiler industry.
Carcass traits are important economic traits in animal production and directly reflect meat production performance of animals, such as carcass weight, half-bore weight, full-bore weight, leg muscle weight, breast muscle weight, wing weight and the like, so that the carcass traits are always hot spots in animal breeding production. On the basis of meat quality, further improvement of the dressing percentage of the broiler chickens is the target of broiler chicken breeding.
non-ATPase3 (protosome 26S subenit, non-ATPase3, PSMD 3) is one of proteasome 26S subunit, a member of non-ATPase family gene, and a member of 19S regulatory complex in 26S proteasome, and the recent research shows that PSMD3 has important function in mitochondria, and the interaction with mitochondria-encoded protein extremely remarkably influences the normal expression of mitochondrial function. At present, the specific role of the PSMD3 gene in birds is rarely reported, and particularly, the PSMD3 gene is less researched in chickens.
Disclosure of Invention
The invention aims to provide a PSMD3 gene molecular marker related to chicken skin color traits and carcass traits and application thereof, so as to solve the problems in the prior art, discover the SNP site of the PSMD3 gene, analyze the SNP site in association with the chicken skin color traits and carcass traits and provide a new SNP molecular marker for MAS.
In order to achieve the purpose, the invention provides the following scheme:
the invention provides a molecular marker related to chicken skin color traits and carcass traits, wherein the molecular marker is positioned on a chicken PSMD3 gene, and the accession number of the PSMD3 gene is GeneID:426133; the nucleotide sequence of the molecular marker is shown as SEQ ID NO. 1 and comprises mutation sites SNP1-SNP3; SNP1 is located in NC _052558.1: G4304798A > G; SNP2 is located in NC _052558.1: G4304889G > A; SNP3 is located in NC _052558.1: g4304903C > T.
Preferably, SNP1 presents genotypes AA, AG and GG; SNP2 has genotypes GG, AG and AA; SNP3 presents genotypes CC, CT and TT.
The invention also provides a method for identifying the chicken skin color character and/or carcass character by using the molecular marker, which comprises the following steps:
(1) Taking chicken genome DNA to be detected as a template, and amplifying a gene segment containing the molecular marker by using a primer to obtain an amplification product; the primers comprise an upstream primer shown as SEQ ID NO. 2 and a downstream primer shown as SEQ ID NO. 3;
(2) Sequencing the amplification product, and detecting the genotype of the corresponding single nucleotide polymorphism site, wherein the dressing percentage of AA and AG genotype individuals of the mutation site SNP1 is higher than that of GG genotype individuals; the abdominal skin yellowness value of the GG genotype individual of the mutation site SNP2 is higher than that of the AG genotype individual; the dressing percentage of CC and CT genotype individuals of the SNP3 mutation site is higher than that of TT genotype individuals.
Preferably, the amplification reaction system is: template DNA 2. Mu.L, 2 XEs Taq MasterMix (Dye) 25. Mu.L, upstream primer 2. Mu.L, downstream primer 2. Mu.L, and ddH 2 O 19μL。
Preferably, the amplification reaction procedure is: pre-denaturation at 94 ℃ for 2min; denaturation at 94 ℃ for 30s, annealing at 60 ℃ for 30s, extension at 72 ℃ for 30s, and 32 cycles; final extension at 72 ℃ for 3min; storing at 4 deg.C.
Preferably, the skin color trait is abdominal skin yellowness value and the carcass trait is slaughter rate.
The invention also provides application of the molecular marker in breeding of the chicken skin color character and carcass character.
The invention discloses the following technical effects:
according to the invention, by analyzing the PSMD3 gene, the gene is found to have a plurality of SNP sites which are obviously related to the chicken skin color character and carcass character, and simultaneously, the gene is found through experimental verification: molecular marker NC — 052558.1: there was a significant correlation between the G4304798A > G locus and dressing percentage (p < 0.05), molecular marker NC _052558.1: G4304889G > a site has a significant correlation with abdominal skin yellowness values (p < 0.05), molecular marker NC — 052558.1: there was a significant correlation between the g4304903C > T locus and the dressing percentage (p < 0.05). The 3 molecular markers can accurately identify the skin color traits and carcass traits of the chickens, and provide new SNP molecular markers for MAS, thereby providing scientific basis for the breeding of the chickens.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings needed in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and it is obvious for those skilled in the art to obtain other drawings without creative efforts.
FIG. 1 is a schematic diagram showing the position of the PSMD3 primer pair and the length of the product;
FIG. 2 is a diagram showing SNP site genotyping of PSMD3 gene.
Detailed Description
Reference will now be made in detail to various exemplary embodiments of the present invention, examples of which are illustrated in the accompanying drawings, wherein methods, unless otherwise specified, are generally performed using conventional methods, and reagents, unless otherwise specified, are generally commercially available or formulated using conventional methods. This detailed description is not to be taken in a limiting sense, but is to be understood as a more detailed description of certain aspects, features and embodiments of the invention.
It is to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. Further, for numerical ranges in this disclosure, it is understood that each intervening value, between the upper and lower limit of that range, is also specifically disclosed. Every intervening value, to the extent any stated or intervening value in a stated range, and every other stated or intervening value in that stated range, is encompassed within the invention. The upper and lower limits of these smaller ranges may independently be included or excluded in the range.
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. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention. All documents mentioned in this specification are incorporated by reference herein for the purpose of disclosing and describing the methods and/or materials associated with the documents. In case of conflict with any incorporated document, the present specification will control.
It will be apparent to those skilled in the art that various modifications and variations can be made in the specific embodiments of the present disclosure without departing from the scope or spirit of the disclosure. Other embodiments will be apparent to those skilled in the art from consideration of the specification. The description and examples are intended to be illustrative only.
As used herein, the terms "comprising," "including," "having," "containing," and the like are open-ended terms that mean including, but not limited to.
In the following description, all methods involved are conventional in the art unless otherwise specified. All the materials referred to are those which are commercially available from the public unless otherwise specified.
Example 1
1. Materials and methods
1.1 animal samples
The animals tested were 409 80-day-old Jiangfeng spotted-brown chickens (purchased from Jiangfeng industries, ltd., guangzhou city). By collecting 2mL of subcutaneous venous blood, the sample was stored at-80 ℃ and used as a DNA extraction sample. Skin color traits and carcass traits such as subcutaneous fat thickness, intramuscular fat width, full bore weight, half bore weight, abdominal fat weight, leg weight, pectoral muscle rate, leg ratio, abdominal fat rate, half bore rate, full bore rate, and slaughter rate were also recorded.
1.2 Primary reagents
Blood sample DNA extraction kit (brand: OMEGA; cat # D3392; feiyang bioengineering, guangzhou), 2 XEs Taq MasterMix (Dye) (brand: kangshi; cat # CW0690M; kangshi Biotechnology, inc.), DNA marker (brand: novozan; cat # MD101; jiangsu Novozan Biotechnology, inc.), high purity low electroosmosis agarose (brand: tokyo; cat # TSJ001; beijing Strobiological, inc.).
1.3 Experimental methods
1.3.1 primer design
According to the sequence of red jungle (Gallus) PSMD3 gene published by NCBI (National Center for Biotechnology Information Search database) (Gene ID: 426133), primers were designed using NCBI's Primer-BLAST tool, and Primer synthesis service was provided by Guangzhou Ongke Biotechnology, inc. The information about the primer sequences is shown in Table 1, and the pairing positions of the primers on the PSMD3 gene are shown in FIG. 1.
TABLE 1 PCR amplification primer sequences
1.3.2 blood sample DNA extraction
And (4) extracting the DNA of the blood sample by referring to an operation manual of the blood sample DNA extraction kit.
1.3.3 PCR amplification of PSMD3 Gene sequences
The genomic DNA of the 409 individual blood samples was used as a template, and the following reaction system was followed: template DNA 2. Mu.L, 2 XEs Taq MasterMix (Dye) 25. Mu.L, upstream primer 2. Mu.L, downstream primer 2. Mu.L, ddH 2 O 19μL。
Reaction procedure: pre-denaturation at 94 ℃ for 2min; denaturation at 94 ℃ for 30s, annealing at 60 ℃ for 30s, extension at 72 ℃ for 30s, and 32 cycles; final extension at 72 ℃ for 3min; storing at 4 ℃. PCR products were subjected to Sanger sequencing by Gene technology, inc., tianyihui, guangzhou.
1.3.4 SNP determination and genotyping
Analysis of the sequence peak patterns was performed on the Sanger sequencing results of the PCR products using the SeqMan tool of DNAstar software to determine potential SNP sites, and sequencing data for each sample was aligned by this tool for genotyping.
1.3.5 genotype correlation analysis with skin color trait and carcass trait
And carrying out correlation analysis on the skin color character and carcass character data of the individual corresponding to the SNP locus and the genotype by adopting an SAS 9.4 GLM program package.
2 results
2.1 PSMD3 gene sequence PCR amplification and SNP screening
Selecting blood sample DNA of 409 individuals of the domestic chicken as a template to carry out PCR amplification, carrying out Sanger sequencing on an obtained PCR product (the nucleotide sequence is shown as SEQ ID NO: 1), carrying out comparison analysis on a peak diagram after sequencing, detecting 3 SNP sites in total, and associating with the chicken skin color character and carcass character, wherein the three SNP sites are respectively as follows: NC _052558.1: G4304798A > G, NC _052558.1: G4304889G > A and NC _052558.1: g4304903C > T, as shown in FIG. 2.
SEQ ID NO:1：
CACAACGTCATCAAGACGGGTAGGAGGTGTGGTTGGGGGCCGTGCCACCACCAGAGCCTCGGTGGGAGGCTGCCACCCTGTGCTGCATGGTGGGAGGCTCCGTTCCTGTTTCCCCTCCCCCCCCAGGTGTCCGCATGATCAGCCTCTCCTACTCCCGCATCTCCCTGGCTGACATTGCCCAGAAGCTGCAGCTGGACAGCCCCGAGGATGCGGAGTTCATTGTTGCCAAGGTAGCTGTGCAGGAAGCCTCGTCCCTCTGCCTCCCGGGGCTGCCCACCACTCCTGCCAGGCCTCTCCTGCCCCGCCCCCAACGCGCTCCCTTCCCAGGGCTGCTGATTTCAGGCTGTGCTGCTCTGGGGGTGCTGAGGGCACCAACTCCCATCTCTGTGCACTCTCTGAGCCCACCGCTCCCAGCGCCTTCCTTTGCTCTGAACTGCTGCCTCCACCCTGGGAGGTGGTGGTGCCCTGACCACGCACCCCGTGGCTCGCAGGCCATCCGGGATGGTGTGATTGAGGCCAGCATTAACCACGAGAAGGGCTACGTGCAGTCAAAAGAGATGATTGACATCTACTCGACCCGGGAGCCCCAGCTGGCGTTCCACCAGCGCATCTCCTTCTGCCTCGACATCCACAACATGTCAGTCAAGGTGAGCACAGCACAGCTCTGTGAGGACCCGGGGGGGGGGTTGGGGGGGCACGGGGATGGCGCAGAGGGAGGAAGCGGTTCGGCTCAGGCTGCAGTAGAGCTGCTGTGAGTGAAAGGACAGCAGAG。
Note: the underlined positions in SEQ ID NO. 1 are SNP sites.
2.2 Correlation analysis of SNP site of PSMD3 gene sequence with skin color trait and carcass trait
Correlation analysis was performed for the above 3 SNP sites with skin color traits and carcass traits (thick subcutaneous fat, wide intramuscular fat, full bore weight, half bore weight, abdominal fat weight, leg weight, pectoral muscle rate, leg ratio, abdominal fat rate, half bore rate, full bore rate, slaughter rate, live body weight, shin length, shin girth and carcass cloaca skin color, shoulder skin color, hip skin color, abdominal skin color, chest leg skin color, shin skin color and abdominal fat color yellowness value). ( Note: in tables 2-4, the different shoulder marks of a and b indicate that the difference is significant, and P is less than 0.05 )
As shown in table 2, the results show that NC _052558.1: G4304798A > G locus has a significant correlation with the dressing percentage (p < 0.05), wherein the dressing percentage of AA and AG genotype individuals is significantly higher than that of GG genotype individuals (p < 0.05);
as shown in table 3, NC _052558.1: the G4304889G > a locus had a significant correlation with abdominal skin yellowness values (p < 0.05). The abdominal skin yellowness value of the GG genotype individuals is obviously higher than that of the AG genotype individuals (p < 0.05), and the AA genotype does not have obvious difference with the AG and GG genotype individuals (p > 0.05).
As shown in table 4, NC _052558.1: the g4304903C > T locus had a significant correlation with the dressing percentage values (p < 0.05). Wherein the dressing percentage of CC and CT genotype individuals is obviously higher than that of TT genotype individuals (p < 0.05).
In addition to abdominal skin yellowness values and dressing percentage, the association of 3 SNP sites with the remaining skin color trait and carcass trait did not reach a significant level (p > 0.05).
Table 2 nc _052558.1: G4304798A > G site is associated with carcass traits
Table 3 nc _052558.1: G4304889G > A locus is associated with skin color trait
Table 4 nc_052558.1: g4304903C > T locus is associated with carcass traits
The above-described embodiments are only intended to illustrate the preferred embodiments of the present invention, and not to limit the scope of the present invention, and various modifications and improvements made to the technical solution of the present invention by those skilled in the art without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.
Claims (7)
1. The molecular marker related to the chicken skin color character and carcass character is characterized in that the molecular marker is positioned on a chicken PSMD3 gene, and the accession number of the PSMD3 gene is GeneID 426133; the nucleotide sequence of the molecular marker is shown as SEQ ID NO. 1 and comprises mutation sites SNP1-SNP3; SNP1 is located in NC _052558.1: G4304798A > G; SNP2 is located in NC _052558.1: G4304889G > A; SNP3 is located in NC _052558.1: g4304903C > T.
2. The molecular marker according to claim 1, wherein SNP1 presents genotypes AA, AG and GG; SNP2 has genotypes GG, AG and AA; SNP3 presents genotypes CC, CT and TT.
3. A method for identifying a skin color trait and/or a carcass trait in a chicken using the molecular marker of claim 1, comprising the steps of:
(1) Taking genome DNA of a chicken to be detected as a template, and amplifying a gene fragment containing the molecular marker in claim 1 by using a primer to obtain an amplification product; the primers comprise an upstream primer shown as SEQ ID NO. 2 and a downstream primer shown as SEQ ID NO. 3;
(2) Sequencing the amplified product, and detecting the genotype of the corresponding single nucleotide polymorphism site of the molecular marker, wherein the dressing percentage of AA and AG genotype individuals of the mutation site SNP1 is higher than that of GG genotype individuals; the abdominal skin yellowness value of the GG genotype individual of the mutation site SNP2 is higher than that of the AG genotype individual; the dressing percentage of CC and CT genotype individuals of the SNP3 mutation site is higher than that of TT genotype individuals.
4. The method of claim 3, wherein the amplification reaction system is: template DNA 2. Mu.L, 2 XEs Taq MasterMix (Dye) 25. Mu.L, upstream primer 2. Mu.L, downstream primer 2. Mu.L, and ddH 2 O 19μL。
5. The method of claim 3, wherein the amplification reaction is performed by: pre-denaturation at 94 ℃ for 2min; denaturation at 94 ℃ for 30s, annealing at 60 ℃ for 30s, extension at 72 ℃ for 30s, and 32 cycles; final extension at 72 ℃ for 3min; storing at 4 ℃.
6. The method of claim 3, wherein the skin color trait is abdominal skin yellowness value and the carcass trait is slaughter rate.
7. Use of a molecular marker as claimed in any one of claims 1 to 2 in the breeding of chicken skin tone traits and carcass traits.
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| CN116790767A (en) * | 2023-07-21 | 2023-09-22 | 华南农业大学 | TYR gene molecular marker related to chicken skin blackness and application thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN116790767A (en) * | 2023-07-21 | 2023-09-22 | 华南农业大学 | TYR gene molecular marker related to chicken skin blackness and application thereof |
| CN116790767B (en) * | 2023-07-21 | 2024-02-20 | 华南农业大学 | TYR gene molecular marker related to chicken skin blackness and application thereof |
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| CN115896299B (en) | 2023-10-13 |
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