CN115851982B - Long-distance molecular genetic marker for chickens and application thereof - Google Patents
Long-distance molecular genetic marker for chickens and application thereof Download PDFInfo
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- CN115851982B CN115851982B CN202211476514.5A CN202211476514A CN115851982B CN 115851982 B CN115851982 B CN 115851982B CN 202211476514 A CN202211476514 A CN 202211476514A CN 115851982 B CN115851982 B CN 115851982B
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- 241000287828 Gallus gallus Species 0.000 title claims abstract description 60
- 235000013330 chicken meat Nutrition 0.000 title claims abstract description 48
- 230000002068 genetic effect Effects 0.000 title claims abstract description 26
- 239000003550 marker Substances 0.000 title claims abstract description 21
- 238000009395 breeding Methods 0.000 claims abstract description 35
- 230000001488 breeding effect Effects 0.000 claims abstract description 34
- 238000012408 PCR amplification Methods 0.000 claims abstract description 5
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 5
- 238000012163 sequencing technique Methods 0.000 claims abstract description 4
- 101150073528 PTN gene Proteins 0.000 claims description 6
- 108700028369 Alleles Proteins 0.000 claims description 4
- 238000011144 upstream manufacturing Methods 0.000 claims description 4
- 239000002773 nucleotide Substances 0.000 claims description 2
- 125000003729 nucleotide group Chemical group 0.000 claims description 2
- 239000013589 supplement Substances 0.000 claims description 2
- 239000003147 molecular marker Substances 0.000 abstract description 4
- 244000144977 poultry Species 0.000 abstract description 4
- 235000013594 poultry meat Nutrition 0.000 abstract description 4
- 230000000694 effects Effects 0.000 abstract description 3
- 230000007547 defect Effects 0.000 abstract description 2
- 238000010353 genetic engineering Methods 0.000 abstract description 2
- 239000003102 growth factor Substances 0.000 abstract description 2
- 230000004983 pleiotropic effect Effects 0.000 abstract description 2
- 238000000034 method Methods 0.000 description 11
- 238000003205 genotyping method Methods 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 3
- 239000009705 sanhuang Substances 0.000 description 3
- 230000003321 amplification Effects 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 238000002844 melting Methods 0.000 description 2
- 230000008018 melting Effects 0.000 description 2
- 238000003199 nucleic acid amplification method Methods 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 238000000137 annealing Methods 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 230000008030 elimination Effects 0.000 description 1
- 238000003379 elimination reaction Methods 0.000 description 1
- 244000144972 livestock Species 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 235000013372 meat Nutrition 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
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- 238000004806 packaging method and process Methods 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 230000000644 propagated effect Effects 0.000 description 1
- 125000006239 protecting group Chemical group 0.000 description 1
- 238000010187 selection method Methods 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
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Abstract
The invention provides a long-distance molecular genetic marker of chicken and application thereof, and relates to the technical field of poultry genetic engineering breeding. The long-distance molecular genetic marker of the chicken is a 63366 locus SNP (63366:C > T) of chicken pleiotropic growth factor (PT N) gene intron 4. The molecular genetic marker genotype is obtained by a primer F:5'-CCAAACCTCTTCTGCGTCTC-3' and R:5'-GCAGAGCATTGTGTCAGGAA-3' PCR amplification sequencing. The invention overcomes the defects of the prior art, provides a long-distance molecular genetic marker for chickens, and utilizes the molecular marker to carry out molecular marker assisted selection on chicken distance length, so that early seed selection on the distance length can be realized, the breeding cost is reduced, the breeding efficiency and effect are improved, and the breeding genetic progress is accelerated.
Description
Technical Field
The invention relates to the technical field of poultry genetic engineering breeding, in particular to a long-distance molecular genetic marker of chickens and application thereof.
Background
The distance of the chickens is located at the rear side of the shin, which is the second sex characteristic of the roosters, and the distance length of the roosters increases with the increase of the age of the day, while the distance of the hens is basically degraded, so the distance length character can be regarded as a restriction character.
Although the distance length has no significant correlation with chicken productivity and meat quality, the distance length is an important packaging trait for roosters, consumers like and preferentially purchase roosters longer in distance length, indicating that the distance length can increase the economic value of the chickens.
When new varieties of broilers are cultivated, increasing the distance length of roosters is often used as one of breeding targets. Since the distance length characteristics of a cock are generally easy to measure after 3 months of age, current selection of distance length is generally carried out after 3 months of age of a breeder hen, an individual selection method is adopted for breeder selection, and breeder selection is carried out only on the cock, and the breeder is not carried out due to no distance length phenotype value. This method has the following drawbacks: 1. late seed selection, namely late elimination and raising cost increase, resulting in high breeding cost; 2. the genetic progress of the selection is slow because the selection of genetic potential cannot be performed on hens.
Disclosure of Invention
Aiming at the defects of the prior art, the invention provides a long-distance molecular genetic marker for chickens and application thereof. By utilizing modern biotechnology and a candidate gene analysis method, the SNP at 63366 locus of chicken pleiotropic growth factor (PTN) gene intron 4, namely PTN (63366:C > T), is found to be obviously related to chicken distance length, wherein the chicken distance length relationship of different genotypes is CC type > CT type > TT type. The mutation can be used as a molecular marker of chicken distance long characters, is used for auxiliary selection of chicken distance long and small molecular markers, realizes early seed selection of the chicken distance long characters, reduces cost, improves breeding efficiency and effect, and accelerates cultivation of new long-distance broiler strains or varieties.
In order to achieve the above object, the technical scheme of the present invention is realized by the following technical scheme:
A long-distance molecule genetic marker of chicken, which is a 63366-locus SNP of intron 4 of a PTN gene.
Preferably, the SNP genotype at 63366 locus of the intron 4 of the PTN gene is obtained by PCR amplification and sequencing by the following primers, and the amplified nucleotide sequence is shown as SEQ ID NO:1, and SNP genotyping is shown in FIG. 1.
Upstream primer F:5'-CCAAACCTCTTCTGCGTCTC-3';
The downstream primer R:5'-GCAGAGCATTGTGTCAGGAA-3'.
The long-distance dominant allele (C) is applied to breeding of chickens, and long-distance chicken breeds can be obtained.
Preferably, the method of application comprises the following steps:
(1) Pure breeding of strains, chickling wearing wing numbers, registering pedigrees, conventional feeding management and relevant properties according to breeding targets;
(2) 6 weeks old, according to breeding targets, carrying out seed selection on other target breeding characters such as weight and the like;
(3) Carrying out genotype identification on the long-distance molecular genetic marker loci of each chicken at 7-8 weeks of age;
(4) Selecting long-distance genotype (CC type) individuals for both the cock and the hen according to the gene identification result in 9-10 weeks;
(5) And when 17 weeks of age, measuring the distance length of the selected and reserved breeder cocks, selecting individuals with larger reserved distance length, and breeding the breeder cocks step by step according to the steps to obtain long-distance chicken strains or breeds.
Preferably, in the step (4), when the number of the long-range genotype individuals selected by the cock and the hen is insufficient, the individuals carrying the long-range dominant allele (C) in the selected portion are selected to supplement the population number.
The invention provides a chicken long-distance molecular genetic marker and application, which have the advantages compared with the prior art:
(1) The molecular genetic marker determined by the invention can assist in selecting chicken breeding, so that the early breeding of hens is realized, and the unnecessary breeding hens are eliminated early, thereby reducing the breeding cost and the breeding cost;
(2) The molecular genetic marker can select the hen which does not show the long-distance character through auxiliary selection, and improves the accuracy and efficiency of seed selection, thereby accelerating the genetic progress of seed selection and accelerating the cultivation of new chicken strains or varieties of long-distance broilers.
Drawings
FIG. 1 is a graph showing the genotyping results of the present invention.
Detailed Description
For the purpose of making the objects, technical solutions and advantages of the embodiments of the present invention more apparent, the technical solutions in the embodiments of the present invention will be clearly and completely described in the following in conjunction with the embodiments of the present invention, and it is apparent that the described embodiments are some embodiments of the present invention, but 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.
Example 1:
The Anhui nan Sanhuang chicken is local genetic resource in China and is listed in the "Chinese livestock and poultry genetic resource and poultry Zhi". The variety belongs to a meat-egg dual-purpose variety, and because of the preference of consumers, the chicken pitch is selected, the growth speed of the chicken pitch is accelerated, and the variety meets the market demand, namely, the Anhui nan Sanhuang chicken is taken as an example, and the long-pitch strain is bred:
1. breeding targets: basically maintains the growth speed of the existing chickens and increases the growth speed of the distance, namely increases the distance length, and breeds to obtain a new long-distance Anhui nan Sanhuang chicken strain (Q2 line).
2. The breeding process comprises the following steps:
(1) Q2 is pure breeding: 10000 chickens are obtained through pure breeding, the male and female anus-turning method is used for identification, about 5000 chickens are reserved, about 2000 chickens are reserved, each chicken wears wing numbers, and a pedigree is registered;
(2) Body weight selection of Q2: through conventional feeding management, weighing the whole group at 6 weeks of age, recording the weight of each chicken, eliminating about 4000 individual hens with oversized weight and undersized weight and centered weight, and about 1500 hens;
(3) Blood samples were collected from individual hens and hens at 8 weeks of age, DNA was extracted, and SNP at 63366 locus of intron 4 of the PTN gene was genotyped according to the following typing method:
① PCR amplification primers were designed for SNP at 63366 locus of intron 4 of the PTN gene, wherein:
The upstream primer sequence is F:5'-CCAAACCTCTTCTGCGTCTC-3';
the downstream primer sequence is R:5'-GCAGAGCATTGTGTCAGGAA-3'.
② PCR amplification was performed using chicken DNA templates and primers:
The amplification step includes: a PCR reaction system and a PCR reaction program;
The PCR reaction system comprises 1 mu L of template DNA, 1 mu L of upstream primer, 1 mu L of downstream primer, 2X HieffTMPCRMasterMix mu L of ddH 2 O7 mu L of downstream primer and 1 mu L of PCR reaction system;
The PCR reaction program comprises the following steps: the first step of initial denaturation at 94 ℃ for 5min, the second step of melting at 94 ℃ for 30s, the third step of annealing at 55 ℃ for 30s, the fourth step of extension at 72 ℃ for 1min, wherein the number of melting to extension cycles is 35, the fifth step of final extension at 72 ℃ for 10min, and the sixth step of preservation at 4 ℃;
③ Sequencing and genotyping the PCR products:
After the PCR products are sequenced by Sanger, all individuals are subjected to genotyping aiming at a marker SNP as shown in figure 1, wherein the genotypes comprise three types of CC, CT and TT, the CC genotype is a long-distance genotype, and the C is a long-distance dominant allele;
Obtaining 399 CC-type cocks, 270 CT-type cocks and 811 TT-type cocks according to the typing; CC-type hen 1253, CT-type hen 555, TT-type hen 2172;
(4) After genotyping, at 9 weeks of age, selecting genotypes, 200 roosters with good growth from 399 CC-type roosters, and eliminating the rest roosters; selecting 1100 hens with good growth and development from 1253 CC hens, selecting 300 hens with good growth and development from 555 CT hens, and eliminating the rest hens;
(5) Measuring the distance length of the cock at 17 weeks of age, and selecting about 100 long-distance individuals;
(6) The Q2 line is continuously and purely propagated in the next generation, and the breeding method of the previous generation is repeated until CT type and TT type individuals are completely eliminated.
3. The breeding effect is as follows:
after 2 generations of breeding, the distance measurement results of the 20-week-old cock are shown in the following table 1 compared with the protected group (as a control group):
TABLE 1 analysis of chicken distance Length Effect Using PTN (63366: C > T) markers
| Group of | Q2 is | Protective group (control group) |
| Breeding method with long-distance (mm) of the seed | 5.44±1.80 | 5.37±1.75 |
| First generation breeding distance is long (mm) | 5.71±1.68a | 5.34±1.72b |
| Second generation breeding distance (mm) | 5.97±1.55A | 5.42±1.83B |
Wherein the superscript letter a, b represents P <0.05; a and B represent P <0.01.
As shown in the table 1, the distance length of the cock of 20 weeks age is increased by 0.53mm and the amplification reaches 9.74% by two generations of molecular marker assisted selection; and the distance length of the group-keeping group of the control group is basically unchanged. Meanwhile, the variation coefficient of the Q2 series distance length is reduced from generation to generation, which shows that the uniformity of the distance length is continuously improved. If the continuous breeding of multiple generations is carried out, the long-distance characteristic becomes more and more obvious.
It is noted that relational terms such as first and second, and the like are used solely to distinguish one entity or action from another entity or action without necessarily requiring or implying any actual such relationship or order between such entities or actions. Moreover, the terms "comprises," "comprising," or any other variation thereof, are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements does not include only those elements but may include other elements not expressly listed or inherent to such process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising one … …" does not exclude the presence of other like elements in a process, method, article, or apparatus that comprises the element.
The above embodiments are only for illustrating the technical solution of the present invention, and are not limiting; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit and scope of the technical solutions of the embodiments of the present invention.
Claims (4)
1. The application of the long-distance molecular genetic marker of the chicken is characterized in that the long-distance molecular genetic marker of the chicken is applied to chicken breeding to obtain long-distance chicken seeds, the genetic marker is 63366 locus SNP of PTN gene intron 4, and the SNP locus is positioned in SEQ ID NO:1, the polymorphism at position 198 is C/T.
2. Use of a long range molecular genetic marker of chicken according to claim 1, characterized in that: the SNP genotype of 63366 locus of the PTN gene intron 4 is obtained by PCR amplification and sequencing by the following primers, and the amplified nucleotide sequence is shown as SEQ ID NO:1 is shown in the specification;
Upstream primer F:5'-CCAAACCTCTTCTGCGTCTC-3';
The downstream primer R:5'-GCAGAGCATTGTGTCAGGAA-3'.
3. Use of a long range molecular genetic marker for chickens according to claim 1, wherein the manner of use comprises the steps of:
(1) Pure breeding of strains, chickling wearing wing numbers, registering pedigrees, conventional feeding management and relevant properties according to breeding targets;
(2) 6 weeks old, according to breeding targets, carrying out seed selection on other target breeding characters such as weight and the like;
(3) Carrying out genotype identification on the genetic marker loci of the long-distance character molecules of each chicken in 7-8 weeks;
(4) According to the gene identification result, the rooster and hen select long-distance genotype, namely CC type individual;
(5) And when 17 weeks of age, measuring the distance length of the selected and reserved breeder cocks, selecting individuals with larger reserved distance length, and breeding the breeder cocks step by step according to the steps to obtain long-distance chicken strains or breeds.
4. Use of a long-range molecular genetic marker for chickens according to claim 3, wherein: in the step (4), when the number of the selected long-distance genotype individuals of the cock and the hen is insufficient, the selected individuals carrying the long-distance dominant allele C can be selected to supplement the population number.
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| EP1027608B1 (en) * | 1998-07-30 | 2010-01-20 | Odyssey Thera, Inc. | Protein fragment complementation assays |
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| CN111647663B (en) * | 2020-06-18 | 2022-07-01 | 安徽农业大学 | Molecular genetic marker for chicken walking number character and application |
| CN111910008B (en) * | 2020-08-21 | 2022-05-31 | 云南农业大学 | Molecular marker related to chicken growth and development and application thereof |
| CN114908176B (en) * | 2022-05-18 | 2022-12-09 | 华南农业大学 | Molecular marker related to chicken carcass and growth traits and application thereof |
| CN115305290A (en) * | 2022-08-30 | 2022-11-08 | 西北农林科技大学 | Chicken liquid chip and application thereof |
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