CN115125308A - SNP molecular marker for genetically improving chicken carcass traits - Google Patents
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Abstract
The invention discloses an SNP molecular marker for genetically improving chicken carcass traits. The invention obtains an SNP molecular marker for genetically improving the chicken carcass traits from a chicken ribonucleoprotein large subunit38 gene (RPL38), and also provides a primer and a kit for detecting the SNP molecular marker. The SNP molecular marker can be used for molecular marker-assisted breeding of chicken carcass traits, has the advantages of low cost and high accuracy, and is beneficial to development of broiler genetic breeding work.
Description
Technical Field
The invention belongs to the technical field of molecular marker breeding. In particular to an SNP molecular marker for genetically improving the carcass trait of chicken.
Background
The carcass traits are one of the most important economic traits of the broiler chickens, and include the indexes such as pre-slaughter live weight, shin length, chest width, half-bore weight, breast weight, leg weight, wing weight, shin-paw weight, small intestine length and the like, and besides the feeding mode has different influences on the carcass traits of the broiler chickens, genetic factors also play a vital role.
Single Nucleotide Polymorphism (SNP) is a molecular breeding technique commonly used in molecular Marker Assisted Selection (MAS), which is widely present in biological populations and can be accurately and stably inherited to offspring. SNP, a stable DNA molecular marker, has been widely used in large-scale breeding work of various animals including chickens. The breeding worker effectively promotes the development of the genetic breeding work of the broilers in China through the related molecular genetic markers established by the research on the SNP. With the development of SNP markers, these markers have been used for screening functional genes for various animal economic traits based on the fine localization of genes, and a large number of excellent genes have been developed and utilized.
The chicken large subunit of ribosomal protein 38 gene (ribosomal protein large subbunit 38, RPL38) is located in the cytoplasm, belongs to the L38E family of ribosomal proteins, and is located on the 18 th chromosome of chicken. In the study of human diseases and plants, the RPL38 gene was once used as a housekeeping gene, and it was considered to be stably expressed in all cells. However, more and more researches show that ribosomal proteins are involved in various ribose in vitro functions such as cell proliferation, cell differentiation, apoptosis, cell development regulation, malignant transformation of normal cells, DNA repair and drug resistance, besides being involved in ribosome composition and influencing protein translation. It has also been found that the RPL38 gene can control the expression of the gene through the internal ribosome entry site, even through binding to the ribosome to control the translation, thereby regulating the development of animal embryo. However, no report on RPL38 has been found in poultry breeding work, and it is not clear whether RPL38 can affect poultry carcass traits.
Disclosure of Invention
The invention aims to enrich SNP molecular markers related to chicken carcass traits, provide an SNP molecular marker for genetically improving chicken carcass traits and promote the development of broiler genetic breeding work in China.
The first purpose of the invention is to provide a SNP molecular marker for genetically improving the carcass traits of chickens.
The second purpose of the invention is to provide a detection primer of the SNP locus.
The third purpose of the invention is to provide a detection kit for detecting the SNP molecular marker.
The fourth purpose of the invention is to provide the application of the SNP molecular marker, the detection primer or the detection kit in the molecular marker-assisted breeding of the chicken carcass traits.
The fifth object of the present invention is to provide a method for screening chicken individuals having good carcass traits.
A sixth object of the present invention is to provide a method for genetically improving the carcass trait of chickens.
The above purpose of the invention is realized by the following technical scheme:
the SNP molecular marker for genetically improving the chicken carcass traits is obtained by carrying out single nucleotide polymorphism detection on the chicken ribonucleoprotein large subunit38 gene (RPL38, positioned on chromosome 18) and analyzing the correlation between different genotypes and the chicken carcass traits, and a scientific basis is provided for molecular breeding of auxiliary chicken carcass traits.
The invention provides an SNP molecular marker for genetically improving the carcass traits of chickens, wherein the nucleotide sequence of the SNP molecular marker is shown as SEQ ID NO. 1; the 216 th site of the nucleotide sequence shown in SEQ ID NO.1 is a SNP site: c > T; the carcass traits of chicken individuals marked with the SNP molecular marker as CC genotypes are obviously higher than those of chicken individuals marked with CT or TT genotypes.
In particular, the carcass traits comprise pre-slaughter live weight, shin length, chest width, chest depth, body lean length, half bore weight, breast weight, leg weight, wing weight, shin-paw weight and/or small intestine length.
Based on the SNP molecular marker provided by the invention, a person skilled in the art can design and obtain a detection primer for amplifying the SNP locus provided by the invention by using a conventional primer design method, and detect the genotype of the SNP locus in a chicken individual by using the designed detection primer for molecular marker-assisted breeding of chicken carcass traits. Therefore, the invention also applies for protecting the detection primer for detecting the SNP locus.
Specifically, the detection primer is designed based on a sequence shown in SEQ ID NO.1, and an amplification sequence of the detection primer contains the SNP locus.
The invention also provides a detection kit for detecting the SNP molecular marker, and the kit contains the detection primer.
Besides the primers, the kit also contains reagents required by PCR amplification.
In view of the fact that the SNP molecular marker is significantly related to the chicken carcass trait, the invention also applies to protect the application of the SNP molecular marker, the detection primer or the detection kit in the molecular marker-assisted breeding of the chicken carcass trait.
The invention also provides a method for screening chicken individuals with good carcass traits, which comprises the following steps: the detection primer of the SNP locus is utilized to amplify the genome DNA of the individual chicken to be detected and carry out the genotyping of the SNP locus, and the chicken individual with CC genotype as the genotyping result is the chicken individual with good carcass traits.
The invention also provides a genetic improvement method for the chicken carcass traits, which comprises the following steps: the strain cultured by the method for improving the carcass traits of the chickens is used as a breeding hen, and the frequency of the dominant allele of the SNP molecular marker loci is improved generation by generation through breeding, so that the carcass traits of the offspring chickens are improved.
The invention has the following beneficial effects:
the SNP molecular marker for genetically improving the chicken carcass trait is obtained by carrying out single nucleotide polymorphism detection on the chicken large subunit ribonucleoprotein 38 gene (RPL38) and analyzing the correlation between different genotypes and the chicken carcass trait. In addition, the invention also provides a primer and a kit for detecting the SNP molecular marker. The SNP molecular marker, the primer and the kit for detecting the SNP marker can be used for screening chicken individuals with good carcass traits, improving the carcass traits of the chicken and carrying out genetic improvement. The SNP molecular marker can be used for molecular marker-assisted breeding of chicken carcass traits, has the advantages of low cost and high accuracy, and is beneficial to development of broiler genetic breeding work.
Drawings
FIG. 1 shows SNP sites according to the present invention: c > T typing scheme.
Detailed Description
The invention is further described with reference to the drawings and the following detailed description, which are not intended to limit the invention in any way. Reagents, methods and apparatus used in the present invention are conventional in the art unless otherwise indicated.
Unless otherwise indicated, reagents and materials used in the following examples are commercially available.
Example 1 PCR amplification of the Chicken RPL38 Gene
1. Animal material
325 chickens of 91-day-old apricot blossom chickens and recessive white rock F2 generation resources are selected, 1.2mL of infrawing venous blood is collected by using a vacuum anticoagulation tube, and the collected blood is stored in a low-temperature refrigerator at the temperature of minus 80 ℃ for subsequent DNA extraction.
The live weight, carcass weight, half-bore weight, full-bore weight, breast weight, leg weight, wing weight, shin-paw weight, small intestine length, chest width, chest depth, body slant length, shin length, small intestine length, etc. of the chicken before slaughtering are measured.
2. Blood DNA extraction
All individual chicken DNA extraction operations were performed according to the instructions of the DNA extraction Kit (NRBC Blood DNA Kit, OMEGA, D0715), and the extracted DNA was subjected to concentration and purity detection and then stored in a low-temperature refrigerator at-20 ℃ for subsequent PCR amplification.
3. Primer design
The invention designs an amplification primer based on an RPL38 Gene sequence (Gene ID: 771528) provided by NCBI official website, the information of the primer is shown in Table 1, the primer is synthesized by Guangzhou branch of Biotechnology, Inc. of Beijing Ongke, and the length of a PCR product is 912 bp.
TABLE 1 PCR amplification primer information for RPL38 gene
4. PCR amplification of chicken RPL38 Gene
The present invention uses 2 × Taq MasterMix (Dye) (kang century, CW0682) reagent for PCR amplification of RPL38 gene, the PCR reaction system is shown in Table 2, the PCR reaction program is operated according to the instruction of 2 × Taq MasterMix (Dye), and the annealing temperature is 58 ℃.
TABLE 2 PCR amplification System
The PCR product was sent to Guangzhou division, Kyoho, for Sangge sequencing, and the nucleotide sequence of the PCR product was as follows (SEQ ID NO. 1):
CCAAAGCCAAGTTGCAGCTGGTGGGAAGCTATTTGCAAATATTACCCT GCCTGTGACTTCGTAAGTCTTTGACAAGCTAAAAGATGTCGTTCCGCTAAA GCTCTGCGACACCTGACAAGTGAATCACGCTTAAGCTCAAGTTTTCTATTG TTGCGAATACAAAGACAATTATAATTGAAAAGCTGGGCTGTCTTGTCTCGGT ATTTAACAACGTGCGAGGAGTTTTCCTTTATTTTTTCCTGTATTTGGATGGA GCTGTCATGGGAAGAGGTGCTTTAATTCTGGAAATGTTTTGTTAACTCAGC GGTGTGGGCGAGAGAGGGATAGCTCCGTGCACACTGCTATTGTAGTTGCTG ACAGCTGCTTACAAAGCATTTCAATACCTTTCAACTCGTTCTAGATTTTGCT GCAGCTCTACTGCTTGTAATGCTGAGCTGCCAGGGCTGTGCAGTGGGAAAG GAGAGCGACTGACCACAGCTCTGAGCAAGCTGCTTAGATAAAACAGCCCT GCGTGGCTATAAAGGATAGGAAGCACTGCTTGCAGCCATGCAGAGTTGAGC AGACCGATTACAGGGGTCGGTGCTGCTATTCAGATGCAAAGGATTACACGA GGAAATCGGTCCAGCATGCACTGCTATGCTCTATTGAGAGTTTGTGATGCTG TGGGCTTCTGATGCTACTTGTAAATATCTGCTATGAGGCAAAAAGATCACGA TAAAGCTGTTTGCTTTGAGAGACCTGAATAGTGTCTGGGAGGCAGCATGCC GAACCCTGTGCAGCTGGCCATCAGCACTGTAGAGAAAGTGCTCAGTTTGA ATTTTCCTCAGCACGTGAGGGGTTGCAGTTGTCCTCCATGCTGAGGGGAAA TAGGAGAACTGGAGTGGGAAATGGTTCGTAAAAGGGACTGCGTGG
this sequence is identical to that shown in > Chromosome 18: 9734285-9735196.
Sequencing result detection finds that the 9734500 site (namely the 216 th site of the sequence shown in SEQ ID NO.1) of the chicken in the RPL38 gene is an SNP mutation site 9734500: c > T.
Example 2 genotyping of SNP sites, Gene frequency calculation, and genotype frequency calculation
The invention utilizes DNAstar software to carry out sequencing on SNP site 9734500 of the Sanger sequencing result: peak patterns of C > T were analyzed, and the SNP sites: the typing scheme of C > T is shown in FIG. 1, and comprises three genotypes of CC, CT and TT. The invention also provides a method for detecting the SNP locus: genotype frequency and allele frequency calculations were performed for the C > T typing results, which are shown in table 3:
table 3 SNP position 9734500: genotype frequency and allele frequency results for C > T
Example 3 SNP site 9734500: association analysis of C > T and carcass traits
The invention uses a mixed linear model in SPSS software (version 24.0) to analyze the correlation between different genotypes of SNP loci and different phenotypic characters of apricot blossom chicken X recessive white rock chicken F2 generation resource groups.
The analytical model is as follows: yijklm ═ μ + Gi + Sj + Hk + fl + eijklm; wherein Yijklm represents a trait phenotype value of an individual, μ represents a population mean value, Gi represents a genotype effect value (i ═ 3) of a marker, Sj represents a gender effect value, Hk represents a batch effect value, fl represents a family effect value, and eijklm represents a random error value; in the operation process, Gi, Sj and Hk are used as fixed factors, and fl is used as a random factor; multiple comparisons were performed using the Borferroin method, with p <0.05 indicating a significant level.
The results of genotyping 325 apricot blossom chickens x recessive white rock F2 generation flock and statistical analysis of carcass traits such as carcass weight, half bore weight, full bore weight, breast weight, leg weight, wing weight, shin and paw weight, small intestine length, chest width, chest depth, body slant length, shin length, and small intestine length of chickens before slaughter are shown in table 4:
table 4 SNP position 9734500: correlation analysis result of C > T and carcass traits
Note: mean ± SE, Mean ± sem; n, the number of individuals; the difference between the lower case letters on the same row indicates significant difference (p < 0.05), and the absence of letter notation indicates no significant difference (p > 0.05).
From the above results, the carcass traits of chicken with SNP molecular marker CC genotype, specifically live weight before slaughter, shin length, chest width, chest depth, body slant length, half-bore weight, breast weight, leg weight, wing weight, shin-paw weight and small intestine length, were significantly higher than those of CT or TT genotype chicken.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
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Claims (10)
1. An SNP molecular marker for genetically improving the carcass traits of chickens, which is characterized in that the nucleotide sequence of the SNP molecular marker is shown as SEQ ID NO. 1.
2. The SNP molecular marker according to claim 1, wherein the 216 th position of the nucleotide sequence shown in SEQ ID No.1 is the SNP site: c > T.
3. The SNP molecular marker according to claim 2, wherein the SNP molecular marker is characterized in that the carcass traits of chicken individuals with CC genotype are significantly higher than those of chicken individuals with CT or TT genotype.
4. SNP molecular marker according to claim 3, characterized in that the carcass trait comprises pre-slaughter live weight, shin length, chest width, chest depth, oblique length, semi-bore weight, chest weight, leg weight, wing weight, shin-paw weight and/or small intestine length.
5. The primer for detecting SNP site according to claim 2, wherein the primer for detecting SNP site is designed based on the sequence shown in SEQ ID No.1, and the amplified sequence contains the SNP site according to claim 2.
6. The primer for detecting SNP sites according to claim 5, wherein the nucleotide sequence of the upstream primer is shown as SEQ ID No.2, and the nucleotide sequence of the downstream primer is shown as SEQ ID No. 3.
7. A detection kit for detecting the SNP molecular marker according to any one of claims 1 to 4, wherein the kit contains the detection primer according to claim 5 or 6.
8. Use of the SNP molecular marker according to any one of claims 1 to 4, the detection primer according to claim 5 or 6 or the detection kit according to claim 7 in molecular marker-assisted breeding of chicken carcass traits.
9. A method for screening chicken individuals with good carcass traits is characterized in that a detection primer for detecting the SNP locus in claim 2 is used for amplifying the genome DNA of a chicken individual to be detected and carrying out genotyping, and the chicken individuals with CC genotypes as the genotyping results are the chicken individuals with good carcass traits.
10. A method for genetically improving the carcass trait of a chicken, comprising breeding a strain bred by the method of claim 9 as a breeding hen to increase the frequency of the dominant allele of the SNP molecular marker locus of claim 2 generation by generation, thereby improving the carcass trait of a offspring chicken.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109182542A (en) * | 2018-10-17 | 2019-01-11 | 华南农业大学 | One kind SNP marker relevant to chicken Carcass Traits and its application |
| CN113913538A (en) * | 2021-11-30 | 2022-01-11 | 华南农业大学 | SNP molecular marker related to chicken carcass traits and application |
| CN114150070A (en) * | 2020-09-08 | 2022-03-08 | 河南农业大学 | SNP molecular marker related to chicken growth and slaughter traits, detection primer, kit and breeding method |
| CN114438231A (en) * | 2022-03-10 | 2022-05-06 | 华南农业大学 | CRELD1 gene molecular marker related to chicken carcass traits and application |
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2022
- 2022-06-07 CN CN202210636903.3A patent/CN115125308A/en active Pending
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|---|---|---|---|---|
| CN109182542A (en) * | 2018-10-17 | 2019-01-11 | 华南农业大学 | One kind SNP marker relevant to chicken Carcass Traits and its application |
| CN114150070A (en) * | 2020-09-08 | 2022-03-08 | 河南农业大学 | SNP molecular marker related to chicken growth and slaughter traits, detection primer, kit and breeding method |
| CN113913538A (en) * | 2021-11-30 | 2022-01-11 | 华南农业大学 | SNP molecular marker related to chicken carcass traits and application |
| CN114438231A (en) * | 2022-03-10 | 2022-05-06 | 华南农业大学 | CRELD1 gene molecular marker related to chicken carcass traits and application |
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| 无: "rs735463976", ENSEMBL, 20 December 2021 (2021-12-20), pages 1 - 4 * |
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