CN111004851B - Detection method and application of correlation between VIPR1 gene and cock body quality and slaughter trait - Google Patents
Detection method and application of correlation between VIPR1 gene and cock body quality and slaughter trait Download PDFInfo
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Abstract
The invention discloses a detection method and application based on correlation between a VIPR-1 gene C1853921T locus and cock body quality and slaughter traits. The invention uses VIPR-1 gene C1853921T locus as a candidate marker to detect the polymorphism of the locus in local cock and analyze the relation between the polymorphism and the body quality and slaughter traits of different ages of the local cock, so as to provide a basis for breeding local cock breeders and marker-assisted selection. Compared with the existing slaughtering and breeding, the breeding method has the advantages of low cost, simple and convenient operation and accurate result. The method is simple and easy to implement, has strong repeatability and can be carried out in a common laboratory.
Description
Technical Field
The invention relates to the field of biological genes and zoology, in particular to a detection method based on correlation between a VIPR1 gene C1853921T locus and cock body quality and slaughter traits and application thereof.
Background
In vivo and in vitro expression experiments of poultry reproductive endocrine regulation mechanism and vasoactive intestinal peptide type I receptor (VIPR 1) gene and correlation between VIPR1 gene polymorphism and reproductive traits prove that the VIPR1 gene is involved in the regulation of reproductive behaviors (Kansaku et al, 2001; You et al, 2001; Zhou et al, 2008; Zhoumin et al, 2011). The avian VIPR-1 polymorphism and the correlation between the polymorphism and the reproductive performance of the female poultry have been studied in a large quantity. The applicant selects 12 polymorphic loci from a chicken VIPR1 gene as a candidate gene of body quality, performs genotype detection on 586 Ningdu yellow hens and analyzes the correlation with the body quality at 77-day-old, 84-day-old and 91-day-old, and finds that the C1853921T locus is only obviously correlated with the body quality at 84-day-old (P < 0.05). However, whether the locus is related to the quality of the male poultry body and the slaughtering traits is not reported. The VIPR-1 gene C1853921T locus is used as a candidate marker to detect the polymorphism of the locus in local cock and analyze the relation between the polymorphism and the body quality and slaughter traits of different ages of the local cock, so as to provide a basis for breeding local cock breeders and marker-assisted selection.
Disclosure of Invention
In order to overcome the defects, the invention provides a method for detecting the correlation between the polymorphism of the C1853921T locus in the VIPR1 gene of the chicken and the body quality and slaughter traits of cocks of different ages and using the correlation and the method as a means for breeding and marking local breeding cocks.
In order to achieve the purpose of the invention, the invention adopts the technical scheme that: a detection method based on the correlation of a VIPR1 gene C1853921T locus and cock body quality and slaughter traits is characterized by comprising the following steps:
(1) extracting the genome DNA of the chicken species to be detected;
(2) amplifying 203bp fragments on the upstream and downstream of the C1853921T locus of the VIPR1 gene by adopting a PCR reaction;
(3) carrying out RFLP typing on the obtained PCR product;
(4) the SAS 9.0GLM program was used to statistically analyze the correlation of the different genotypes of VIPR1 gene locus C1853921T with cock body quality and slaughter traits.
Step (1), collecting 1-2 mL of blood by using a parafin vein, anticoagulating with 2% EDTA, and storing at-20 ℃ for later use. The blood samples were extracted with genomic DNA by conventional phenol/chloroform extraction and diluted to 100 ng/. mu.L for use.
Step (2) downloading a chicken VIPR1 genome sequence (GenBank accession number: NM-0010975230) from NCBI, and designing an upstream primer and a downstream primer by using Genetool software, wherein the primer sequences are as follows: 5'-agaggaacgcagccagtg-3' and 5'-cccacctaacataaaagctcaac-3', and amplifying 203bp fragments upstream and downstream of the C1853921T site of the VIPR-1 gene. The primers were synthesized by Hainan Okangke BioLimited.
And (3) performing RFLP typing on the detected PCR product. The enzyme digestion reaction system is as follows: PCR product6.5. mu.L of the product, 0.3. mu.L of endonuclease BsuR I, 1.0. mu.L of 10 XBuffer buffer, ddH2O2.2. mu.L, left overnight in an incubator at 37 ℃. Detecting the enzyme digestion product by 2 percent agarose gel electrophoresis. The gel imaging system takes pictures and judges the genotype according to the banding pattern.
Performing statistical analysis by adopting an SAS 9.0GLM program in the step (4), and constructing a model as follows: yij ═ μ + Gi + eij. Wherein YIj is a trait phenotypic value, mu is an overall mean value of the trait, Gi is a genotype effect value, and eij is a random residual effect.
Application of the C1853921T locus of the VIPR1 gene in detecting correlation of cock body quality and slaughter traits.
The application specifically comprises the steps of analyzing the relationship between the genotype and the body quality and slaughter traits of cocks of different ages by taking the gene locus C1853921T of the breeding cock VIPR1 as a candidate marker, and breeding local breeding cocks.
Compared with the prior art, the invention adopts a method for detecting the correlation between the polymorphism of the C1853921T locus contained in the VIPR1 gene of the chicken variety and the body quality and slaughter traits of the cock in different weeks, and uses the correlation and the method as a means for breeding and marking the chicken variety. Compared with the existing slaughtering and breeding, the breeding method has the advantages of low cost, simple and convenient operation and accurate result. The method is simple and easy to implement, has strong repeatability and can be carried out in a common laboratory.
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FIG. 1 shows the cleavage at the C1853921T site of VIPR1 gene; wherein: m is DS2000 DNA Marker1, 2: type TC; 3. 4, 5, 7: type TT; 6: type CC.
Detailed Description
The following detailed description further describes the present invention for the purpose of illustrating the technical solutions and objects of the present invention.
1 materials and methods
1.1 test materials, index determination and sample Collection
The test chicken flock is provided by south Jiangxi teacher science and technology Limited, the feeding time is 4 months-8 months in 2018, the number of the young chicken is 700 feathers, the feeding mode is a breeding cock cage-breeding mode, unified immunization is carried out according to a conventional immunization program of broiler breeders, and the rest is carried out feeding management according to a conventional method. The wing size was worn on the first day of birth and the foot size was worn on week 5. Body mass was measured every week from the birth of test chickens, and slaughtered after raising to 16 weeks of age, and measured. The index measurements include carcass mass, semi-bore mass, full-bore mass, semi-lateral pectoral mass, and semi-lateral leg mass. Remove death, escape, and obvious errors and duplicate data, and finally obtain 499 test cocks. The index measurement is carried out according to the method specified in NY/T823-2004 "poultry Performance noun terminology and metrics statistics method". Collecting blood 1-2 mL in a infrawing vein, anticoagulating with 2% EDTA, and storing at-20 ℃ for later use. The blood samples were extracted with genomic DNA by conventional phenol/chloroform extraction and diluted to 100 ng/. mu.L for use.
1.2 primer design and Synthesis
The genomic sequence of chicken VIPR1 (GenBank accession No.: NM-0010975230) was downloaded from NCBI, and the upstream and downstream primers were designed using Genetool software, primer sequences: 5'-agaggaacgcagccagtg-3' and 5'-cccacctaacataaaagctcaac-3', and amplifying 203bp fragments upstream and downstream of the C1853921T site of the VIPR-1 gene. The primers were synthesized by Hainan Okangke BioLimited.
The PCR reaction system was (10. mu.L): 2 XPCR mix 5. mu.L, upstream and downstream primers 0.2. mu.L each, DNA template 0.6. mu.L, ddH2O 4. mu.L. The PCR reaction program is: pre-denaturation at 95 ℃ for 3 min; denaturation at 95 ℃ for 30s, annealing at 58 ℃ for 30s, extension at 72 ℃ for 30s, and 35 cycles; post extension was carried out at 72 ℃ for 5 min. The PCR product was electrophoresed through 1% agarose gel to determine if the fragment size was as expected. And typing the PCR product after detection by using RFLP. The enzyme digestion reaction system is as follows: 6.5. mu.L of PCR product, 0.3. mu.L of endonuclease BsuR I, 1.0. mu.L of 10 XBuffer buffer, ddH2O2.2. mu.L, left overnight in an incubator at 37 ℃. Detecting the enzyme digestion product by 2 percent agarose gel electrophoresis. The gel imaging system takes pictures and judges the genotype according to the banding pattern.
1.3 statistical analysis
As the observed groups have the same genetic background and are all 16w cocks, and the cocks are raised under the same raising standard, the association analysis between the marker and the growth and slaughter traits is statistically analyzed by adopting the SAS 9.0GLM program, and the model is constructed as follows: yij ═ μ + Gi + eij. Wherein YIj is a trait phenotypic value, mu is an overall mean value of the trait, Gi is a genotype effect value, and eij is a random residual effect.
2 results and analysis
The correlation of different genotypes of the VIPR1 gene locus C1853921T with growth and slaughter traits is shown in Table 1. As can be seen from table 1, there were significant (P <0.05) or very significant (P <0.01) differences in 17 personality growth, slaughter traits among the 22 traits tested, 4w to 16w body mass (P <0.05) or very significant (P <0.01 higher than those of TT and TC type individuals) for CC type individuals, and carcass mass, half bore mass, full bore mass, and half side leg mass were very significantly higher than those of TT and TC type individuals (P <0.01) for CC type individuals. Unrelated to other 5 personality (P > 0.05).
TABLE 1 correlation of site C1853921T genotype with growth and slaughter traits of caged Ningdu yellow rooster
Based on the obtained correlation analysis results, the breeding method can be used for screening and breeding the genotype chicken species with excellent data.
The foregoing shows and describes the general principles and broad features of the present invention and advantages thereof. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, which are described in the specification and illustrated only to illustrate the principle of the present invention, but that various changes and modifications may be made therein without departing from the spirit and scope of the present invention, which fall within the scope of the invention as claimed. The scope of the invention is defined by the appended claims and equivalents thereof.
Claims (4)
- A method for detecting correlation between VIPR1 gene and cock body quality and slaughter traits, which is characterized by comprising the following steps:(1) extracting the genome DNA of the chicken species to be detected;(2) amplifying 203bp fragments on the upstream and downstream of the C1853921T locus of the VIPR-1 gene by adopting a PCR reaction;(3) carrying out RFLP typing on the obtained PCR product;(4) adopting SAS 9.0GLM program to carry out statistical analysis on the correlation between different genotypes of VIPR1 gene locus C1853921T and cock body quality and slaughter traits;the primer sequence used for PCR amplification in the step (2): 5'-agaggaacgcagccagtg-3' for F, 5'-cccacctaacataaaagctcaac-3' for R;the chicken breeds are Ningdu yellow chickens;the cock body mass and slaughter shape refer to carcass mass, half-bore mass, full-bore mass, half-side leg mass.
- 2. The method for detecting correlation between VIPR1 gene and cock body quality and slaughter traits as claimed in claim 1, wherein the statistical analysis is performed by SAS 9.0GLM program in step (4), and the model is constructed as follows: yij ═ μ + Gi + eij, where Yij is the phenotypic value of the trait, μ is the overall mean of the trait, Gi is the genotypic effect value, and eij is the random residual effect.
- 3. The application of a detection primer of a C1853921T locus of VIPR1 gene in detecting correlation of cock body quality and slaughter traits, wherein the chicken is Ningdu yellow chicken, cock body quality and slaughter shape refer to carcass quality, half-bore quality, full-bore quality and half-side leg quality, and the sequence of the detection primer is as follows: 5'-agaggaacgcagccagtg-3' for F and 5'-cccacctaacataaaagctcaac-3' for R.
- 4. Use according to claim 3, characterized in that: the application specifically comprises the steps of analyzing the relationship between the genotype, the body quality of different ages and slaughter traits of local chicken breeders by taking the VIPR1 gene locus C1853921T of the chicken as a candidate marker, and breeding local chicken breeders.
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| CN104561279A (en) * | 2014-12-22 | 2015-04-29 | 江苏省家禽科学研究所 | Method of improving quality of chicken semen, primer used for method, kit and using method of kit |
| CN104946776A (en) * | 2015-07-09 | 2015-09-30 | 马韫韬 | Chicken FABP4 gene molecular genetic marker related to good chicken slaughtering character and application of chicken FABP4 gene molecular genetic marker |
| CN105441560A (en) * | 2015-12-29 | 2016-03-30 | 云南农业大学 | Chicken body size trait breeding molecular marker IGF-1R (insulin-like growth factor-1 receptor) gene and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104561279A (en) * | 2014-12-22 | 2015-04-29 | 江苏省家禽科学研究所 | Method of improving quality of chicken semen, primer used for method, kit and using method of kit |
| CN104946776A (en) * | 2015-07-09 | 2015-09-30 | 马韫韬 | Chicken FABP4 gene molecular genetic marker related to good chicken slaughtering character and application of chicken FABP4 gene molecular genetic marker |
| CN105441560A (en) * | 2015-12-29 | 2016-03-30 | 云南农业大学 | Chicken body size trait breeding molecular marker IGF-1R (insulin-like growth factor-1 receptor) gene and application thereof |
Non-Patent Citations (1)
| Title |
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| Identification and optimization of small molecule antagonists of vasoactive intestinal peptide receptor-1 (VIPR1);Lalgudi S.Harikrishnan等;《Bioorganic & Medicinal Chemistry Letters》;20120315;第22卷(第6期);第2287-2290页 * |
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