A method for breeding Sansui duck for growth and meat characteristics

Abstract

The present invention discloses a method for breeding Sansui ducks for growth and meat traits, which is bred by combining a 70-day-old comprehensive index selection method with a SNP marker-assisted breeding method. The 70-day-old comprehensive index selection method comprises the following steps: taking the 70-day-old body weight and the 70-day-old daily average weight gain as the breeding target traits, constructing an individual comprehensive selection index formula, calculating the comprehensive index value, selecting excellent individuals according to the breeding group size and seed retention rate, and forming a family; the Sansui duck growth trait SNP marker-assisted breeding method comprises the following steps: carrying out genotyping for SNP sites that are significantly associated with the 70-day-old body weight and the 70-day-old daily average weight gain, and carrying out assisted breeding according to the site genotype and the SNP combination haplotype. The present invention can accurately and quickly improve the growth and meat performance of Sansui ducks and accelerate the genetic improvement progress of Sansui ducks.

Description

Technical Field

[0001] The invention belongs to the technical field of livestock and poultry breeding, and specifically relates to a method for breeding Sansui ducks for growth and meat characteristics. Background Art

[0002] With the improvement of people's living standards, breeding new breeds of low-fat small-sized meat ducks has good market development potential. Excellent meat duck breeds require that the breast and leg meat accounts for about 30% of the total net carcass weight, of which the breast meat rate is not less than 13%. Sansui duck is an important local duck genetic resource in Guizhou Province, with a breeding history of more than 600 years. Sansui duck is a small egg-laying duck breed with the characteristics of early maturity, high egg production, strong vitality, tender meat and unique flavor. Studies have shown that the slaughter rate of Sansui duck is higher than 90%, the breast and leg muscle rate is higher than 30%, and the breast muscle rate is about 15%, which meets the standards of high-quality meat ducks. The sebum rate is 15%, which is lower than that of meat duck breeds and most local duck breeds. The above shows that Sansui duck has good potential for breeding high-quality low-fat meat ducks. However, its scientific research investment in breeding is obviously insufficient. In recent years, it has been seriously invaded by foreign breeds. It is urgent to strengthen its own breeding, improve the genetic performance of the breed, and tap its excellent characteristics. Sansui ducks grow slowly, the heritability of growth and meat traits is low, and the adult weight is 1450-1680 grams. Traditional breeding methods have many selection indicators and progress slowly, and have little effect in the later stages of breeding. In order to improve the genetic quality of Sansui ducks, increase their growth rate and slaughter meat performance, cultivate new high-quality meat Sansui duck strains, optimize target traits and establish effective selection methods. Summary of the invention

[0003] The purpose of the present invention is to provide a method for breeding Sansui ducks for growth and meat characteristics, so as to solve the problems existing in the above-mentioned prior art.

[0004] In order to achieve the above-mentioned purpose, the present invention adopts the following technical solutions:

[0005] A method for breeding Sansui ducks for growth and meat traits is to use a 70-day-old comprehensive index selection method combined with a SNP marker-assisted breeding method for breeding.

[0006] The 70-day-old comprehensive index selection method comprises the following steps:

[0007] Taking the 70-day-old body weight and the 70-day-old daily weight gain as the breeding target traits, we constructed the individual comprehensive selection index formula, calculated the comprehensive index value, and selected excellent individuals and established families according to the breeding group size and seed retention rate.

[0008] The SNP marker-assisted breeding method comprises the following steps:

[0009] Genotyping was carried out on SNP loci that were significantly associated with body weight at 70 days of age and average daily weight gain at 70 days of age, and assisted breeding was carried out based on the locus genotype and SNP combination haplotype.

[0010] Furthermore, the method further comprises the step of performing conventional breeding before the age of 70 days, wherein the step adopts the following method:

[0011] Before 70 days of age, weak individuals or those with genetic defects are eliminated. Between 40 and 70 days of age, body shape and appearance assessment is carried out based on the breed characteristics of Sansui Duck, and individuals that do not meet the breed characteristics are eliminated.

[0012] The present invention firstly carries out body shape and appearance identification before 70 days of age to eliminate individuals that do not meet the breed characteristics and genetic dysplasia, with the purpose of eliminating the influence of foreign blood infiltration in the breeding of Sansui ducks, ensuring the uniformity of the group, and selecting excellent individuals at the same time. Then, through research, it is determined that the target traits for optimization selection are body weight at 70 days of age and average daily weight gain at 70 days of age. By calculating phenotypic and genetic parameters, the selection index coefficient is determined, and a comprehensive selection index formula is constructed. Through individual phenotypic measurement, the comprehensive selection index value is calculated according to the selection index formula, and individual phenotypic selection is carried out.

[0013] The present invention mines the trait-significantly associated SNPs by measuring and analyzing the body weight, body size, slaughter and meat quality of Sansui duck population at 0-22 weeks of age, as well as whole genome sequencing and trait association analysis. The best target trait is determined by calculating parameters such as trait index, phenotypic correlation, genetic correlation, heritability, and the like, and a comprehensive index selection formula is formulated. The inventors have found through a large number of studies that the average daily weight gain at 10 weeks of age is significantly correlated with breast muscle rate, breast muscle shear force, breast meat brightness value, carcass weight, whole eviscerated weight, keel length, chest width, etc., which are indirect selection traits of the above traits.

[0014] Furthermore, the comprehensive selection index formula is:

[0015]

[0016] Where I is the comprehensive index value; T 70 is the individual weight at 70 days of age, AVG is the average weight of the breeding group at 70 days of age; 70 The average daily weight gain of individuals at 70 days of age, It is the average daily weight gain of the breeding group at 70 days of age.

[0017] Furthermore, there are four SNP sites significantly associated with the body weight at 70 days of age and the average daily weight gain at 70 days of age, namely SNP1, SNP2, SNP3 and SNP4, which are located at the 209th, 215th, 219th and 1259th sites from the 5' end of the nucleotide sequence shown in SEQ ID No.1, respectively.

[0018] Furthermore, the SNP1 corresponds to the 114656458 site on the duck chromosome 3 sequence in the GenBank database (NCBI Reference Sequence: NC_051774.1); the genotypes include CC, TC and TT.

[0019] Furthermore, the SNP2 corresponds to the 114656464 site on the duck chromosome 3 sequence in the GenBank database (NCBI Reference Sequence: NC_051774.1); the genotypes of the SNP2 include CC, TC and TT.

[0020] Furthermore, the SNP3 corresponds to the 114656468 site on the duck chromosome 3 sequence in the GenBank database (NCBI Reference Sequence: NC_051774.1); the genotypes of the SNP3 include AA, GA and GG.

[0021] Furthermore, the SNP4 corresponds to the 114657508 site on the duck chromosome 3 sequence in the GenBank database (NCBI Reference Sequence: NC_051774.1), and the genotypes of the SNP4 include TT, CT and CC.

[0022] Furthermore, the SNP marker-assisted breeding method for growth traits of Sansui duck comprises the following steps:

[0023] ① Extracting the genomic DNA of Sansui duck to be tested, performing PCR amplification using a primer pair, and after PCR cycles, amplifying a PCR product with a nucleotide sequence as shown in SEQ ID NO: 1, wherein the nucleotide sequence also includes the four SNP sites SNP1-SNP4;

[0024] ②PCR product purification and sequencing;

[0025] ③ Select individuals whose genotypes of SNP1, SNP2, SNP3 and SNP4 are CC, CC, AA and TT respectively. SNP1, SNP2, SNP3 and SNP4 form a haplotype block. Select individuals whose genotype of the haplotype block is CCAT as breeding ducks and eliminate individuals with other haplotypes.

[0026] Furthermore, after PCR amplification, the PCR product was 1142 bp in length, including positions 151 to 1292 of the SEQ ID NO.1 sequence, corresponding to positions 114656400 to 114657541 on the duck chromosome 3 sequence in the GenBank database (NCBI Reference Sequence: NC_051774.1).

[0027] The advantages and beneficial effects of the present invention are:

[0028] (1) The present invention provides a method for breeding Sansui ducks for growth and meat traits. First, the 70-day-old body weight and the 70-day-old daily average weight gain are used as breeding target traits, a comprehensive selection index formula is constructed, and phenotypic breeding is carried out; secondly, SNP sites that are extremely significantly associated with the two target traits are disclosed, and the dominant genotype or combined haplotype of the SNP site has the optimal growth and meat trait, and genetic marker-assisted breeding is carried out with the SNP site. The present invention establishes an effective Sansui duck breeding selection index formula, and provides a target trait selection index method and a SNP marker-assisted breeding method. The two methods are used in combination to carry out Sansui duck breeding, which can accurately and quickly improve the growth and meat performance of Sansui ducks, realize precise genetic breeding, quickly improve production performance, and accelerate the genetic improvement progress of Sansui ducks. The method is not affected by age, gender, feeding management and other environmental factors, shortens the breeding progress, improves the breeding efficiency, and has important economic value and practical significance for improving the genetic performance of Sansui ducks, reducing costs and increasing efficiency.

[0029] (2) The present invention takes the body weight at 70 days of age and the average daily weight gain at 70 days of age as target traits, optimizes breeding indicators, constructs a selection index, and converts individual phenotypic values ​​into selection index values. By directly selecting the index values, the selection efficiency can be improved, and ultimately the overall market weight and economic benefits of the breeding population can be improved.

[0030] (3) Due to the significant correlation between the traits, the selection and breeding of the two target traits can indirectly improve the group's breast muscle rate, carcass weight, eviscerated weight, body size and other indicators.

[0031] (4) By selecting the genotypes and haplotypes of SNP loci that are significantly associated with the two target traits, body weight at 70 days of age and average daily weight gain at 70 days of age, the influence of the feeding environment can be eliminated, economic traits can be improved from a genetic perspective, and breeding progress can be improved. BRIEF DESCRIPTION OF THE DRAWINGS

[0032] Figure 1 This is the cumulative growth curve of Sansui duck from 0 to 22 weeks of age. The vertical axis in the figure is the weight data of each week of age, and the horizontal axis is the two-week age;

[0033] Figure 2 This is the SNP density map after genome-wide association quality control. DETAILED DESCRIPTION

[0034] The following will be combined with the drawings in the embodiments of the present application to clearly describe the technical solutions in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, rather than all the embodiments. All other embodiments obtained by ordinary technicians in this field based on the embodiments in the present application belong to the scope of protection of this application.

[0035] The experimental methods in the following examples, unless otherwise specified, are all conventional methods, and are performed according to the techniques or conditions described in the literature in the field or according to the product instructions. The materials, reagents, etc. used in the following examples, unless otherwise specified, can all be obtained from commercial channels.

[0036] Embodiment: This embodiment provides a method for breeding Sansui duck for growth and meat characteristics

[0037] 1. Test materials:

[0038] The experiment used 655 Sansui ducks (483 female ducks and 172 male ducks) from the breeding duck farm of Sansui County Qianlishan Food Technology Co., Ltd. Male and female ducks were mixed and raised under the same batch hatching, same feeding conditions and environment.

[0039] 2. Body shape and appearance identification:

[0040] The body shape and appearance assessment was completed at the age of 40-70 days. The assessment was conducted based on feather color and body shape, and other features such as the head, beak, and feet were also checked. Specifically, (1) Feather color. For male ducks, select individuals with brown feathers from the neck to the chest, dark brown feathers on the back, light brown feathers on the abdomen, and individuals with dark green luminous feathers on the neck and tail waist. For female ducks, select individuals with nubile feathers. At the same time, the feathers under the wings of male and female ducks are required to be white and without any other colors. (2) Body shape. The neck is slender, the body is long and wide, the chest is broad and prominent, the sternum is long, the tail is upturned, the body is boat-shaped, and it is about 50 degrees to the ground when walking. (3) Other features. The head is small and the beak is short, the iridescence is brown, the shanks are slender, and the shanks and webbed feet are orange-red. After the body shape and appearance assessment, 287 individuals that did not meet the requirements were eliminated, and 368 individuals remained.

[0041] 3. Production data determination:

[0042] The body weight of the children aged 0-22 weeks was measured every 2 weeks, and the average daily weight gain was calculated and a growth curve was drawn. Figure 1 .from Figure 1 It can be seen that in the first 10 weeks (70 days of age, i.e. Figure 1 The 5-week-old corresponding position is the key growth period of Sansui ducks, and the growth is slow in the later period. The weight at 70 days of age can be used as the target trait for the breeding of Sansui ducks, and early breeding can be carried out.

[0043] After 70 days of age, body size, slaughter and meat quality were measured. The measurement and calculation methods were based on the "Terminology and Statistical Methods of Poultry Production Performance: NY / T823-2004". The correlation analysis results of the average daily weight gain at 70 days of age and body size, slaughter and meat quality are shown in Table 1.

[0044] Table 1 Correlation between average daily weight gain and various traits at 70 days of age

[0045]

[0046] Note: The numbers in the table are the correlation coefficients between the average daily weight gain at 70 days of age and various trait indicators, ** indicates extremely significant correlation (P<0.01), and * indicates significant correlation (P<0.05). As shown in Table 1, the average daily weight gain at 70 days of age is significantly correlated with the slaughter traits of adult Sansui ducks, shear force in meat quality indicators, breast muscle brightness value, and keel length and chest width in body size traits, indicating that selecting the average daily weight gain at 70 days of age as the target trait can indirectly improve the slaughter performance of Sansui ducks.

[0047] Beijing Compson Agricultural Technology Co., Ltd. was commissioned to complete the whole genome deep sequencing of all samples and the 70-day-old body weight (T 70 ) and average daily weight gain (AVG 70 ) GWAS analysis. The SNP density map after quality control is attached Figure 2 .

[0048] 4. Comprehensive selection index calculation and individual selection:

[0049] To introduce the technical solution of the present invention in more detail, the data of 65 samples randomly selected from 368 are taken as an example below.

[0050] According to the formula:

[0051]

[0052] Calculate the comprehensive selection index value.

[0053] The calculation results of the comprehensive index values ​​of the 65 samples are shown in Table 2 below.

[0054] 5. SNP marker-assisted breeding method for growth traits of Sansui duck:

[0055] Genotyping was carried out on SNP loci that were significantly associated with body weight at 70 days of age and average daily weight gain at 70 days of age, and assisted breeding was carried out based on the locus genotype and SNP combination haplotype.

[0056] This example uses direct sequencing of PCR products to determine the locus genotype. The blood sample DNA was extracted using the TIANamp Blood DNA Kit, and the primer sequence was synthesized by Tiangen Biochemical Technology Co., Ltd. PCR reaction system: 1 μL DNA template, 1 μL each of upstream and downstream primers, 10 μL of 2×Taq PCR MasterMix reagent, and ddH2O was added to 20 μL. PCR amplification reaction procedure: ① Pre-denaturation. 95°C 6min, ② Amplification reaction. 95°C denaturation for 30s, 59°C annealing for 30s, 72°C extension for 45s, 35 cycles; finally 72°C extension for 7min. The PCR product was commissioned to Nosai Genomic Research Center Co., Ltd. for sequencing.

[0057] The PCR amplification primer sequence information is as follows:

[0058] Upstream primer F: 5'-CCTTTGGTCTCAGGACCCAG-3'; (as shown in SEQ ID No. 2)

[0059] Downstream primer R: 5'-GGCATGGCTATCCCCAAAGA-3'; (as shown in SEQ ID No. 3).

[0060] The nucleotide sequence of the amplified product is shown in SEQ ID No. 1. The PCR product is 1142 bp in length, including positions 151 to 1292 of the SEQ ID NO. 1 sequence, corresponding to positions 114656400 to 114657541 on the duck chromosome 3 sequence in the GenBank database (NCBI Reference Sequence: NC_051774.1).

[0061] DNAMAN software was used for sequence alignment and genotyping, and the genotype results of each locus of the 65 samples are shown in Table 2. The four loci SNP1-SNP4 in Table 2 correspond to loci 114656458, 114656464, 114656468 and 114657508 on chromosome 3, respectively.

[0062] It should be noted that the SNP markers provided by the present invention can be amplified by the PCR primers provided above and combined with sequencing for genotyping, and typing can also be achieved using other PCR primers or primer combinations, direct deep sequencing, nucleic acid sequence probes and the like. The above methods are all within the technical solution of the present invention and cannot be used as a means to limit the scope of the present invention.

[0063] Table 2 Comprehensive index values ​​and SNP genotypes of 65 samples

[0064]

[0065]

[0066] SPSS22.0 software was used to compare and analyze the body weight at 70 days of age and average daily weight gain at 70 days of age for different genotypes at each site. The results are shown in Table 3.

[0067] Table 3 Comparison of body weight and average daily weight gain at 70 days of age among locus genotypes

[0068]

[0069]

[0070] As shown in Table 3, the dominant genotypes of the above four loci are CC, CC, AA and TT. The results of the significant comparison test showed that the body weight at 70 days of age and the average daily weight gain at 70 days of age were significantly higher than those of other genotypes (P<0.05).

[0071] 6. Individual selection and elimination

[0072] The selection conditions include: (1) selecting individuals with higher index values; (2) selecting the 209th, 215th, 219th and 1259th sites from the 5' end on SEQ ID No. 1, corresponding to the 114656458, 114656464, 114656468 and 114657508 sites on the duck chromosome 3 sequence in the GenBank database (NCBI Reference Sequence: NC_051774.1), and the individuals with genotypes of CC, CC, AA and TT, respectively, that is, selecting individuals with a 4-site combined haplotype of CCAT, and eliminating individuals with other haplotypes.

[0073] According to the selection conditions, population size and breeding plan, a specific selection plan is formulated. The population of this embodiment includes 65 individuals, and the population size is small. The average value of 65 individuals is 1.04, and the standard deviation is 0.26. With the average value-0.5*standard deviation, i.e., 0.91, individuals with index values ​​higher than 0.91 are selected, and individuals with index values ​​lower than 0.91 are eliminated. The eliminated individuals include 24 individuals with sequence numbers 9, 10, 11, 12, 20, 23, 26, 28, 30, 33, 37, 38, 39, 40, 42, 47, 48, 50, 51, 52, 56, 62, 63, and 65. Among the remaining 41 individuals, 10 individuals with sequence numbers 3, 4, 7, 17, 22, 24, 27, 46, 49, and 59 do not meet the haplotype CCAT condition and are eliminated.

[0074] Finally, 31 individuals were selected for breeding. The 70-day-old body weight, 70-day-old daily average weight gain, and 160-day-old live weight, carcass weight, and breast muscle weight of the 31 individuals selected are shown in Table 4. The relevant indicators of the 34 eliminated individuals are shown in Table 5. From Table 4, the weight of the selected individuals at 70 days of age (1275.60±113.83), average daily weight gain at 70 days of age (15.04±3.22), live weight before slaughter at 160 days of age (1390.74±134.39), carcass weight at 160 days of age (1276.31±125.31), and breast muscle weight at 160 days of age (139.66±17.52) were calculated, which were all greater than the weight of the 65 individuals before selection at 70 days of age (1169.41±151.11), average daily weight gain at 70 days of age (12.45±4.12), and live weight before slaughter at 160 days of age (139.66±17.52). The weight of the 34 eliminated individuals at 70 days of age (1072.60±110.71), the average daily weight gain at 70 days of age (10.10±3.4), the live weight before slaughter at 160 days of age (1189.06±137.82), the carcass weight at 160 days of age (1097.01±142.33), and the breast muscle weight at 160 days of age (112.82±19.17) were all greater than the weight of the 34 eliminated individuals at 70 days of age (1072.60±110.71), the average daily weight gain at 70 days of age (10.10±3.4), the live weight before slaughter at 160 days of age (1189.06±137.82), the carcass weight at 160 days of age (1097.01±142.33), and the breast muscle weight at 160 days of age (112.82±19.17). This shows that through this method, individuals with excellent T70 and AVG70 phenotypes can be selected, and the slaughter performance of adult ducks (such as 160 days of age) can be indirectly improved. At the same time, through the selection of SNP loci, the dominant genotypes are aggregated.

[0075] Table 4 Index values ​​of various traits of selected individuals (unit / g)

[0076]

[0077]

[0078] Table 5 Index values ​​of various traits of eliminated individuals (unit / g)

[0079]

[0080]

[0081] In summary, the present invention constructs a comprehensive index formula for the growth and meat traits of Sansui ducks according to the principle of quantitative genetics based on the analysis of a large amount of production data and the calculation of heritage parameters, and carries out phenotypic breeding of growth and meat traits by calculating the comprehensive index value. At the same time, according to the deep sequencing of the whole genome of Sansui ducks and the trait GWAS analysis, the significant SNP genetic loci of the breeding target traits are found, and the genetic marker-assisted selection of growth and meat traits is achieved by selecting the dominant genotype or combined haplotype of the genetic loci. The two are effectively combined to reduce the breeding index, and accurately and quickly improve the growth and meat performance of Sansui ducks, accelerate the genetic improvement of Sansui ducks, tap their growth potential, cultivate new strains of low-fat and high-quality local meat ducks, and have important economic value and scientific significance for the improvement of germplasm resources of local ducks and the reduction of costs and increase of efficiency.

[0082] The embodiments of the present application are described above in conjunction with the accompanying drawings, but the present application is not limited to the above-mentioned specific implementation methods. The above-mentioned specific implementation methods are merely illustrative and not restrictive. Under the guidance of the present application, ordinary technicians in this field can also make many forms without departing from the purpose of the present application and the scope of protection of the claims, all of which are within the protection of the present application.

Claims

1. A method for breeding Sansui duck for meat growth traits, characterized in that: The breeding was carried out by combining the 70-day-old comprehensive index selection method with the SNP marker-assisted breeding method. The 70-day-old comprehensive index selection method comprises the following steps: Taking the 70-day-old body weight and the 70-day-old daily weight gain as the breeding target traits, we constructed the individual comprehensive selection index formula, calculated the comprehensive index value, and selected excellent individuals and established families according to the breeding group size and seed retention rate. The SNP marker-assisted breeding method comprises the following steps: Genotyping was performed on SNP loci that were significantly associated with body weight at 70 days of age and average daily weight gain at 70 days of age, and assisted breeding was carried out based on locus genotypes and SNP combination haplotypes; The comprehensive selection index formula is: , Where I is the comprehensive index value; T 70 is the individual weight at 70 days of age, AVG is the average weight of the breeding group at 70 days of age; 70 The average daily weight gain of individuals at 70 days of age, It is the average value of the average daily weight gain of the breeding group at 70 days of age; There are four SNP sites significantly associated with the body weight at 70 days of age and the average daily weight gain at 70 days of age, namely SNP1, SNP2, SNP3 and SNP4, which are located at the 209th, 215th, 219th and 1259th sites from the 5' end of the nucleotide sequence shown in SEQ ID No.1, respectively; The genotype of SNP1 is CC, TC or TT; the genotype of SNP2 is CC, TC or TT; the genotype of SNP3 is AA, GA or GG; the genotype of SNP4 is TT, CT or CC; the selection conditions include: (1) selecting individuals with higher index values; (2) selecting individuals whose genotypes are CC, CC, AA and TT at the 209th, 215th, 219th and 1259th sites from the 5' end on SEQ ID No.1, respectively, according to the genotypes of the four sites and the site combination haplotypes, that is, selecting individuals whose genotypes are CCAT for the four site combination haplotypes, and eliminating individuals with other haplotypes.

2. The method for breeding Sansui duck for growth and meat characteristics according to claim 1, characterized in that: It also includes a step of conventional selection before 70 days of age, which uses the following methods: Before 70 days of age, weak and genetically defective individuals are eliminated. Between 40 and 70 days of age, body shape and appearance are identified based on the breed characteristics of Sansui Duck, and individuals that do not meet the breed characteristics are eliminated.

3. The method for breeding Sansui duck for growth and meat characteristics according to any one of claims 1 to 2, characterized in that: The SNP marker-assisted breeding method for growth traits of Sansui ducks comprises the following steps: ① Extracting the genomic DNA of Sansui duck to be tested, performing PCR amplification using a primer pair, and after PCR cycles, amplifying a PCR product with a nucleotide sequence as shown in SEQ ID NO: 1, wherein the nucleotide sequence also includes the four SNP sites SNP1-SNP4; ②PCR product purification and sequencing; ③ Select individuals whose genotypes of SNP1, SNP2, SNP3 and SNP4 are CC, CC, AA and TT respectively. SNP1, SNP2, SNP3 and SNP4 form a haplotype block. Select individuals whose genotype of the haplotype block is CCAT as breeding ducks and eliminate individuals with other haplotypes.

4. The method for breeding Sansui duck for growth and meat characteristics according to claim 3, characterized in that: The PCR product has a total length of 1142 bp, including positions 151 to 1292 of the SEQ ID NO.1 sequence, corresponding to positions 114656400 to 114657541 on the duck chromosome 3 sequence with a GenBank database accession number of NC_051774.1.

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