阅读说明：本技术 一种乌骨山羊的提纯繁育方法 (Purification breeding method of black-bone goats ) 是由 杨前平 索效军 熊琪 张年 李晓锋 刘洋 陈明新 陶虎 张凤 于 2020-05-29 设计创作，主要内容包括：本发明涉及山羊繁育技术领域,具体涉及一种乌骨山羊的提纯繁育方法；先通过表型特征构建基础群体,再通过构建TYR模型构建纯种群体,根据家系构成将纯种群体划分为两个品系、并分别采用闭锁群体继代选育和快速繁育以获得稳定的世代,最后将两个稳定世代进行杂交,获得所述乌骨山羊的核心群。本发明通过对乌骨山羊体内的酪氨酸酶(TYR)含量的测定,构建了基础乌骨羊遗传TYR模型,能够促进乌骨山羊的快速繁育,使其优良基因被充分、高效和快速地保护和利用。(The invention relates to the technical field of goat breeding, in particular to a method for purifying and breeding black-bone goats; constructing a basic population through phenotypic characteristics, constructing a pure population through a TYR model, dividing the pure population into two strains according to family composition, respectively adopting locked population subculture breeding and rapid breeding to obtain stable generations, and finally hybridizing the two stable generations to obtain the core population of the black-bone goats. According to the invention, a basic Wugu goat genetic TYR model is constructed by measuring the content of Tyrosinase (TYR) in the Wugu goat body, so that the rapid breeding of the Wugu goat can be promoted, and excellent genes of the Wugu goat can be fully, efficiently and rapidly protected and utilized.)

1. A method for purifying and breeding black-bone goats is characterized by comprising the following steps:

s1: building a basic group: screening by phenotypic characteristics to construct said basal population;

s2: construction of pure seed populations: determining the content of tyrosinase in the plasma of the basic population, constructing a TYR model, eliminating individuals with low or no TYR content and non-UK character, and dividing families to form the pure population;

s3: dividing the pure population into a line 1 and a line 2 according to the family composition; carrying out latch population subculture breeding on the strain 1 to obtain a stable generation of the strain 1; rapidly breeding the strain 2 to obtain a stable generation of the strain 2;

s4: and (3) measuring the combining ability between the stable generation of the line 1 and the stable generation of the line 2, and hybridizing to obtain the core group of the black-bone goats.

2. The method for purifying and breeding the black-bone goats according to claim 1, wherein the phenotypic characteristics comprise: the hair color is pure black, pure white or black and white variegated, the horn of sheep is hornless or the horn is in a semi-spiral shape, the neck of sheep is short and not wrinkled, the temple of sheep is low and small, the chest of sheep is deep and wide, the back is flat and straight, the abdomen is big and not drooping, the body is relatively short, the tail is short and is in a cone shape, the head and the four limbs are not covered enough by the hair, and the hair is thick;

collecting and recording pedigree information, weight and age of each ram and ewe, and establishing a basic group consisting of n families by taking the rams as a unit.

3. The method for purifying and breeding the black-bone goats according to claim 1, wherein the method for constructing the pure population in the step S2 comprises the following steps: evaluating breeding values of individuals in the basal population by using the TYR model, wherein the TYR model is Y ═ Xn × An;

wherein Y is a production trait measured value, n is the number of families, Xn is a corresponding TYR related set value in the nth family, and An is An individual breeding estimation average value in the nth family.

4. The method for purifying and breeding the Wugu goats according to claim 3, wherein the TYR-related set value is obtained by a method comprising the following steps: determining the tyrosinase content in the plasma of all individuals in each of said families and composing a set { Bn }; then Xn is 0 when SDn/{ Bn } Bve <0.05, and 1 when SDn/{ Bn } Bve ≧ 0.05; wherein { Bn } Bve is an average of the values in each of the sets { Bn }; where SDn is the standard deviation of the values in each of the sets { Bn }.

5. The method for purifying and breeding Wugu goats according to claim 4, wherein the breeder population is obtained by selecting the pedigrees corresponding to the rams with the Y value greater than or equal to 60% of the maximum Y value of the rams through genetic evaluation in step 2), and further selecting the ewes with the Y value greater than or equal to 60% of the maximum Y value of the ewes in each pedigree.

6. The method for purifying and breeding the black-bone goats according to claim 5, wherein the breeding method of the line 1 comprises the following steps: and after locked breeding, breeding by applying the TYR model and combining the comprehensive index.

7. The method for purifying and breeding the black-bone goats according to claim 5, wherein the breeding method of the line 2 comprises the following steps: after artificial insemination is adopted, the TYR model is applied to breeding by combining the comprehensive index.

Technical Field

The invention relates to the technical field of breeding of stockings, in particular to a method for purifying and breeding black-bone goats.

Background

The black-bone sheep are black-bone animals (except black-bone chickens) which are determined to have heritable performance, are the only mammals with the black-bone characters found by human beings at present, and are the rare species resources in the world. The black bone membrane and the black meat of the black-bone sheep are the most direct black-bone-shaped manifestations, and the new characters can be stably inherited by the mutation of the line gene. High melanin content is the main cause of the black body expression of black-bone sheep. However, the number of the existing black-bone goats is limited, the rapid propagation of the conventional technology is limited by the number of the groups, and the time interval is long.

Disclosure of Invention

The invention aims to overcome the technical defects, and provides a method for purifying and breeding black-bone goats, which comprises the following steps:

s1: building a basic group: screening by phenotypic characteristics to construct said basal population;

s2: construction of pure seed populations: determining the content of tyrosinase in the plasma of the basic population, constructing a TYR model, eliminating individuals with low or no TYR content and non-UK character, and dividing families to form the pure population;

s3: dividing the pure population into a line 1 and a line 2 according to the family composition; carrying out latch population subculture breeding on the strain 1 to obtain a stable generation of the strain 1; rapidly breeding the strain 2 to obtain a stable generation of the strain 2;

s4: and (3) measuring the combining ability between the stable generation of the line 1 and the stable generation of the line 2, and hybridizing to obtain the core group of the black-bone goats.

Compared with the prior art, the invention has the beneficial effects that:

1. according to the invention, a basic Wugu goat genetic TYR model is constructed by measuring the content of Tyrosinase (TYR) in the Wugu goat body, so that the rapid breeding of the Wugu goat can be promoted, and excellent genes of the Wugu goat can be fully, efficiently and rapidly protected and utilized.

2. The invention further selects the purified population of the TYR as two strains based on the TYR model, and respectively carries out locked population subculture breeding and rapid breeding, and carries out combining ability determination and hybridization to finally reach the core population, the reproductive performance of the obtained core population is improved, and the progress of the TYR, the weight and the wide inheritance of the back drive is stable.

Drawings

FIG. 1 is a schematic diagram of breeding provided by the embodiment of the present invention.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention is described in further detail below with reference to the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

According to the invention, a basic Wugu goat individual genetic TYR model is constructed by measuring the content of Tyrosinase (TYR) in the Wugu goat body, so that the fast breeding of the Wugu goat can be promoted, and excellent genes of the Wugu goat can be fully, efficiently and quickly protected and utilized.

Specifically, referring to fig. 1, a method for purifying and breeding black-bone goats comprises the following steps:

s1: building a basic group: screening through phenotypic characteristics to construct the basic population of the black-bone goats;

s2: construction of pure seed populations: determining the content of tyrosinase in the plasma of the basic population, constructing a TYR model, eliminating individuals with low or no TYR content and non-UK character, and dividing families to form the pure population;

s3: dividing the pure population into a line 1 and a line 2 according to the family composition; carrying out latch population subculture breeding on the strain 1 to obtain a stable generation of the strain 1; rapidly breeding the strain 2 to obtain a stable generation of the strain 2;

s4: and (3) measuring the combining ability between the stable generation of the line 1 and the stable generation of the line 2, and hybridizing to obtain the core group of the black-bone goats.

Selection of basal populations

The phenotype selection characteristics of the black-bone sheep in the basic group building process comprise: the hair color is pure black, pure white or black and white variegated, the horn of sheep is hornless or the horn is in a semi-spiral shape, the neck of sheep is short and not wrinkled, the temple of sheep is low and small, the chest of sheep is deep and wide, the back is flat and straight, the abdomen is big and not drooping, the body is relatively short, the tail is short and is in a cone shape, the head and the four limbs are not covered enough by the hair, and the hair is thick;

652 black-bone sheep are selected from the basic group, wherein 32 ram sheep are aged about 1 year old, the ram sheep are healthy and durable, energetic, moderate in fat condition, the two testicles are the same in size, strong in sexual desire and normal in development. The ewes are 620, the ewes select the ewes to widen, and the breasts of the ewes develop well. Collecting and recording pedigree information, weight and age of each ram and ewe, establishing 32 families by taking the rams as a unit to form a basic group 0 generation, and avoiding the relationship between rams and ewes in each family. The appearance and production performance of the whole ram and ewe are mixed with the conventional requirements and characteristics of the product, and the product is disease-free. And because the black-bone seedlings are oestrous all the year round, the male and female sheep can mate and farrowing all the year round, and the selected male sheep and female sheep are in the breeding period.

Construction of pure seed populations

Specifically, the method for constructing the pure population in the step S2 includes: adopting the basic Wugu sheep individual genetic TYR model to evaluate breeding values of individuals in the basic population, wherein the TYR model is,

Y＝Xn×An

wherein Y is a production trait measured value, n is the number of families, Xn is a corresponding TYR related set value in the nth family, and An is An individual breeding estimation average value in the nth family.

Constructing a Tyrosinase (TYR) standard curve and determining the TYR content of an individual:

by constructing a tyrosinase standard curve (specifically a linear regression curve between a tyrosine light absorption value and a concentration), plasma in an individual is collected, an OD value of the plasma is measured after treatment, the plasma is substituted into the standard curve, the tyrosinase content in each individual is calculated, and the individual with the tyrosinase content lower than 0.1mg/mL is excluded.

Blood is collected by 10mL, heparin sodium is added for centrifugation to prepare blood plasma, and the sample is stored at the temperature of 20 ℃ below zero for later use. Taking 5 test tubes, adding 2mg/m L-dopa solution (L) 1mL and 2.8mL of 0.1mol/L sodium phosphate buffer solution (pH is 7.0)2.8mL respectively, shaking uniformly, preheating in a 25 ℃ constant-temperature water bath kettle for 10min, adding 1mL of Tyrosinase (TYR) standard solution with different concentrations into the test tubes respectively, mixing uniformly immediately, timing accurately, rapidly adding 0.2mL of thiourea-ethanol saturated solution at the time of 2min (complete reaction), mixing uniformly, stopping reaction, taking 0.1mol/L sodium phosphate buffer solution as a blank control tube, and measuring the light absorption value at the wavelength of 475nm in a 722 spectrophotometer. Determination of plasma TYR Activity the tyrosinase standard solution from the above procedure was replaced with 1mL of plasma.

The determination result shows that the mean value of TYR content of the basic population excluding low-content TYR individuals (0.1mg/mL) is 0.25 +/-0.05 mg/mL, and the method has important significance for quickly establishing the basic population. Because, in general, the TYR content of the black-bone goat is not less than 0.1mg/mL, and a higher activity is exhibited. The TYR activity level determines a mechanism of conversion between eumelanin and pheomelanin, and the high-level TYR activity is used for conducting synthesis of the high-level eumelanin, so that the genetic character of the study can be determined, and the basis for constructing a pure population can be obtained through rapid and preliminary screening.

Determination of Xn:

specifically, the method for obtaining the TYR correlation set value Xn includes: determining the tyrosinase content in the plasma of all individuals in each of said families and grouping them into a set { Bn }; when SDn/{ Bn }_Bve<0.05, Xn is 0, when SDn/{ Bn }_BveXn is 1 when not less than 0.05; wherein { Bn }_BveSDn is the standard deviation of the values in each of the sets { Bn }, being the average of the values in each of the sets { Bn }.

Determination of An:

recording three traits of each individual and relatives thereof according to three traits of tyrosinase activity, weight and post-drive width in blood of a basic population, calculating heritability, phenotypic variance and economic weighted value of the three traits and corresponding genetic correlation and phenotypic correlation according to a classical Selection index (Selection index), and making a comprehensive index I, so that after low-content individuals are eliminated by measuring the tyrosinase activity of the individuals, calculating the comprehensive index I in each family, and calculating An average value, wherein the An value is the average value of the comprehensive indexes I of all the individuals in each family.

The quantitative inheritance principle is applied, and according to the inheritance characteristics and economic value of characters, several characters to be selected are integrated into a numerical value which can make the individuals mutually compared, and the numerical value is the selection index. For ease of breed selection, the individual index value of the average number of traits in the flock can be designated as 100, and for other flocks, more individuals than 100 are better, and the formula is:

wherein, the total of three traits (tyrosinase activity, body weight, wide rear drive), W₁h₁ ²+W₂h₂ ²+W₃h₃ ²＝100

Individual phenotype value (Pi) and corresponding average number of flocks

Can be directly calculated from the pasture data. Heritability of traits (h)²) If necessary data are lacked, the method can also be found out from related breeding literature; the breeding or economic importance (Wi) of each trait can be determined empirically by investigation. As for a black-bone sheep individual, the following data are now assumed to be known:

wherein, W₁:W₂:W₃0.51:0.22: 0.27; and, W₁+W₂+W₃＝1

Let the index of the individuals with each trait at the average number of flocks be 100, then:

a₁＝W₁h₁ ²/(W₁h₁ ²+W₂h₂ ²+W₂h₂ ²)×100＝44.07

a₂＝W₂h₂ ²/(W₁h₁ ²+W₂h₂ ²+W₂h₂ ²)×100＝44.07

a₃＝W₂h₂ ²/(W₁h₁ ²+W₂h₂ ²+W₂h₂ ²)×100＝11.86

thus, checking a₁+a₂+a₃When the number is 100, a can be₁、a₂、a₃Respectively substituted into formulas to calculate

I＝44.07/0.25P₁+44.07/85P₂+11.86/24P₃

The formula of comprehensive seed selection index is obtained after simplification, and I is 176.28P₁+0.52P₂+0.49P₃

The selection index I can be calculated by substituting the phenotype value of each character. And calculating the value of I in the nth family to obtain the value of An.

And when the Xn and An values are determined, calculating Y values, sorting the Y values of the rams of the 32 families, and selecting the rams and the corresponding families which account for 60 percent or more of the maximum Y value. Then, further sorting the Y values of the ewes in each family, selecting ewe individuals accounting for 60% of the maximum Y value to construct the family, and eliminating the family if the number of the ewes selected in each family is less than 10. Thus, 15 families, which were the pure population, were obtained, and the results thereof are shown in table 1 below.

TABLE 1

Family system	Ram TYR (mg/mL)	Number of ewes (only)	Average TYR of ewe (mg/mL)
				1#	0.32	25	0.26
2#	0.26	16	0.18
				3#	0.16	13	0.16
4#	0.23	12	0.23
				5#	0.33	11	0.27
6#	0.29	16	0.18
				7#	0.23	25	0.16
8#	0.26	16	0.18
				9#	0.36	12	0.17
10#	0.26	18	0.31
				11#	0.21	20	0.26
12#	0.26	22	0.24
				13#	0.19	11	0.16
14#	0.18	16	0.20
				15#	0.16	12	0.21

Core group construction

Dividing the pure breed group into a strain 1 and a strain 2 according to the family composition, wherein the specific family composition comprises individual TYR content level of ram, number of ewes and average TYR content level of ewes in each family, and is specifically shown in Table 1; the strain 1 is selected by adopting a locked population subculture selection method, and is independently eliminated according to genetic defects to obtain a stable generation of the strain 1; the strain 2 sampling rapid propagation breeding method is characterized in that according to genetic defects, independent elimination is carried out to obtain a stable generation of the strain 2; and (3) determining the combining ability of the generations with stable breeding performance of the line 1 and the line 2, and hybridizing to obtain the core group of the black-bone goats.

And (3) strain 1 breeding: selecting 8 family groups of 3#, 6#, 7#, 8#, 9#, 13#, 14# and 15# in the table 1, wherein the families have no relationship, carrying out locked breeding, adopting the TYR model of the embodiment and breeding by combining the comprehensive index, and the generation interval is 100 d.

And (3) strain 2 breeding: selecting 7 families 1#, 2#, 4#, 5#, 10#, 11# and 12# in the table 1 for rapid breeding, selecting semen of each ram, performing artificial insemination to all ewes in the same family, then adopting the TYR model of the embodiment and combining comprehensive index breeding, establishing the family of the screened herds, and reducing the affinity in the same family as much as possible to obtain 1 generation; the method is adopted for more than 2 generations until the stable black-bone sheep flock with high-level TYR is obtained. The breeding generations of line 1 and line 2 are preferably 2-3 generations. The generation interval is 90 d.

The production performance of the 3 generations of line 1 and the 3 generations of line 2 obtained and the core population after crossing are as follows 2. Table 2 shows that in the breeding process, the TYR performance of the locked population subculture breeding method is relatively consistent, but the breeding performance is reduced; the rapid breeding method has the advantages that the reproductive performance is not degraded, but the genetic progress is unstable; the core group obtained after hybridization is performed by measuring the combining ability of the line 1 and the line 2, the reproductive performance of the obtained core group is improved, and the progress of the TYR, the body weight and the genetic character with wide back drive is stable.

TABLE 2

	Line 1	Line 2	Core group
				Ram size (only)	4	8	12
Average ram TYR (mg/mL)	0.41	0.38	0.37
				Average ram body weight (kg)	78	85	80
Number of ewes (only)	25	31	56
				Average TYR of ewe (mg/mL)	0.39	0.31	0.35
Average weight of ewe (kg)	65	72	68
				Average rear drive width (cm) of ewe	26	35	32

The above-described embodiments of the present invention should not be construed as limiting the scope of the present invention. Any other corresponding changes and modifications made according to the technical idea of the present invention should be included in the protection scope of the claims of the present invention.