CN103609527B - The selection of a kind of blue or green shank and recessive white feather chicken strain - Google Patents
The selection of a kind of blue or green shank and recessive white feather chicken strain Download PDFInfo
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- CN103609527B CN103609527B CN201310589767.8A CN201310589767A CN103609527B CN 103609527 B CN103609527 B CN 103609527B CN 201310589767 A CN201310589767 A CN 201310589767A CN 103609527 B CN103609527 B CN 103609527B
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Abstract
The present invention discloses the selection of a kind of blue or green shank and recessive white feather chicken strain, belongs to molecular breeding technology field. The present invention adopts the genotype in the method qualification dominant Bai Yu site of chicken of molecule assisted Selection, the genotype in this site can be determined fast, producer gene type is the blue or green shank and recessive white feather chicken strain of ii, can effectively avoid surveying and hand over the loaded down with trivial details of seed selection, shorten the generation interval, accelerate breeding process, reduce and cultivate cost. Adopting blue or green shank and recessive its fine and tender taste of white feather chicken strain of the inventive method seed selection, the delicious U.S. of meat, production performance height, both directly can put into production as commercial chicken, it is also possible to carry out supporting application with blue or green shin jute plumage.
Description
Technical field
The present invention is specifically related to the selection of a kind of blue or green shank and recessive white feather chicken strain, belongs to molecular breeding technology field.
Background technology
Along with growth in the living standard, the consumption idea of people is changed into mass type from the scalar type in past. People are also more and more higher to the requirement of chicken meat. The indigenous chicken of China has the good feature of meat, but its speed of growth is relatively slow, and feed conversion rate is low. And though the external fast large-scale broiler chicken speed of growth is fast, feed conversion rate height, its meat is poor. For addressing this problem, large-scale for state's extra income broiler chicken and domestic local high quality meat chicken being hybridized, the meat that can either improve the large-scale broiler chicken of state's extra income also can accelerate the speed of growth and the production performance of local high quality meat chicken. But, due to the impact of the long-term food habits of our people, the chicken of coloured plumages such as Chinese Consumer's preference jute plumage, black plumage, especially Shelter in South China Cities does not accept the white feather chicken of fast-growth. In order to the phenotype of chicken meets the demand of Chinese Consumer's after making hybridization, just need the Recessive Chicken isozygotied, Recessive Chicken production performance height, and the plumage look of coloured plumage after hybridizing, can be kept with China coloured plumage, the breeding for China's high-quality chicken is significant.
Summary of the invention
It is an object of the invention to provide the selection of a kind of blue or green shank and recessive white feather chicken strain.
In order to realize above object, the technical solution adopted in the present invention is:
A selection for blue or green shank and recessive white feather chicken strain, comprises the following steps:
(1) utilizing blue or green shank and recessive white feather chicken as male parent, Bai Laihang chicken hybridizes as female parent, produces F1 generation;
(2) F1 generation selfing obtains F2 generation, the F2 generation blue or green white plumage male and female chicken of shin of selecting and remain;
(3) AS-PCR is utilized to measure the genotype of the F2 generation blue or green white plumage male and female chicken of shin, dominant Bai Yu site of selecting and remain is the individuality of ii genotype, eliminate the individuality that dominant Bai Yu site is II and Ii genotype, obtain the white plumage male and female chicken of blue or green shin that dominant Bai Yu site is ii genotype;
(4) the blue or green shin white plumage male and female chicken selfing that dominant Bai Yu site is ii genotype is got, the method of step (3) is utilized to measure the genotype of self progeny, repeat above-mentioned steps to genetic stability, obtain the blue or green shank and recessive white feather chicken strain that dominant Bai Yu site is ii genotype.
In described step (1), blue or green shank and recessive white feather chicken is the blue or green shin white feather chicken after blue or green shin jute plumage indigenous chicken suddenlys change, and measuring dominant Bai Yu site through the artificial RFLP-PCR method introducing PvuII is ii genotype.
Described step (3) utilize AS-PCR measure the method for F2 generation blue or green shin white plumage male and female chicken genotype, comprise the following steps: taking comprise PMEL17 gene chicken complete genome DNA to be measured as template, increase chicken PMEL17 gene I allelotrope by primer PCR of primer pair P1, and increase chicken PMEL17 gene i allelotrope by primer PCR of primer pair P2; The fragment that amplification obtains is carried out agarose gel electrophoresis, identifies the genotype in the dominant Bai Yu site of chicken according to agarose gel electrophoresis result;
Described primer pair P1 is:
Upstream primer, P1-F:5 '-CCGGCCTCTACTGCCTCAACGTCTCGT-3 ' (as shown in SEQIDNo.1),
Downstream primer, P1-R:5 '-GTGCCCAGAGCTTCGGCGATGAGCGGTG-3 ' (as shown in SEQIDNo.2);
Described primer pair P2 is:
Upstream primer, P2-F:5 '-CCACGCTCACCGTGGGGCTGCTGCTCAT-3 ' (as shown in SEQIDNo.3),
Downstream primer, P2-R:5 '-GAGGGGGTGAGTGCTGTGTTCTC-3 ' (as shown in SEQIDNo.4).
Described pcr amplification reaction system is: 2 �� TaqPCRMasterMix12.5 �� L, ddH2O9.5 �� L, P1-F0.4 �� L, P1-R0.6 �� L, P2-F0.6 �� L, P2-R0.4 �� L, DNA profiling 1.0 �� L.
2 described �� TaqPCRMasterMix contains Taq DNA polymerase, dNTPs, magnesium chloride and PCR reaction buffer, is commercial goods, can purchased from the company such as the raw work in Shanghai, Dalian precious biology, Tyke, Beijing hundred, Beijing Kang Wei.
Described pcr amplification reaction program is: 94 DEG C of denaturation 4min; 94 DEG C of sex change 30s, 64 DEG C of annealing 30s, 72 DEG C extend 15s, 35 circulations; 72 DEG C extend 10min.
The mass concentration of described sepharose is 1.5%.
The voltage of described agarose gel electrophoresis is 120V, and electrophoresis time is 40min.
The described method identifying the dominant white plumage loci gene type of chicken is: II genotype shows as 322bp and 224bp band; Ii genotype shows as 322bp, 224bp and 144bp band; Ii genotype shows as 322bp and 144bp band.
Preferably, the blue or green shank and recessive white feather chicken strain that dominant Bai Yu site step (4) obtained is ii genotype is supporting with blue or green shin yellow pockmarked feather chicken mixing breed, produces coloured plumage broiler chicken.
Described blue or green shin yellow pockmarked feather chicken kind is gu-shi chicken, limit chicken, Huaibei partridge chicken, Huainan Spotted-brown chicken, Wuhua chicken, Chongren fiber crops chicken, Lang Yaji, Lushi chicken, Land of Peach Blossoms chicken, precious jade chicken, camellia chicken etc.
The useful effect of the present invention:
The present invention adopts the genotype in the method qualification dominant Bai Yu site of chicken of molecule assisted Selection, the genotype in this site can be determined fast, producer gene type is the blue or green shank and recessive white feather chicken strain of ii, can effectively avoid surveying and hand over the loaded down with trivial details of seed selection, shorten the generation interval, accelerate breeding process, reduce and cultivate cost. Adopting blue or green shank and recessive its fine and tender taste of white feather chicken strain of the inventive method seed selection, the delicious U.S. of meat, production performance height, both directly can put into production as commercial chicken, it is also possible to carry out supporting application with blue or green shin jute plumage.
For detecting the primer of the white plumage loci gene type of chicken PMEL17 gene dominant in the present invention, it is insert (-CTGGGCACC-) for PMEL17 gene exon10 9bp to design allelotrope I Auele Specific Primer P1-R; For PMEL17 gene exon10 without 9bp insertion design allelotrope i Auele Specific Primer P2-F. Such primer P1-R and P1-F can only increase I allelotrope, and the clip size of amplification is 224bp; Primer P2-R and P2-F can only increase i allelotrope, and the clip size of amplification is 144bp. In addition, owing to the combination of upstream primer P1-F and downstream primer P2-R can also be increased, the clip size of amplification is 322bp, and this amplification is not by the impact of 9bp insertion.
Accompanying drawing explanation
Fig. 1 is the PCR electrophorogram in each sample PMEL17 gene dominant Bai Yu site in the embodiment of the present invention 1;
Note: the electrophoresis band type of swimming lane difference representation DNA MarkerI and genotype ii, ii, Ii, II, II, ii, Ii, ii and Ii from left to right.
Embodiment
The present invention is only described in further detail by following embodiment, but does not form any limitation of the invention.
Embodiment 1
The selection of blue or green shank and recessive white feather chicken strain in the present embodiment, comprises the following steps:
(1) utilizing blue or green shank and recessive white feather chicken in gu-shi chicken colony to make male parent, Bai Laihang chicken hybridizes as female parent, produces F1 generation;
(2) F1 generation selfing obtains F2 generation, the F2 generation blue or green white plumage male and female chicken of shin of selecting and remain;
(3) utilize the method for AS-PCR to measure the genotype of the F2 generation blue or green white plumage male and female chicken of shin, specifically comprise the following steps:
1. the collection of sample
To the F2 blue or green shin white feather chicken wing venous blood collection selected and remain of generation, the antithrombotics that adds 1/3, extracts blood DNA with proteinase-K pathway, DNA sample is stored in 4 DEG C of Refrigerator stores for subsequent use;
2. design of primers
Taking PMEL17 gene order (GenBankAccessionNo.AY636129) as template design primer is to P1 and P2, as follows;
Described primer pair P1 is:
Upstream primer, P1-F:5 '-CCGGCCTCTACTGCCTCAACGTCTCGT-3 ',
Downstream primer, P1-R:5 '-GTGCCCAGAGCTTCGGCGATGAGCGGTG-3 ';
Described primer pair P2 is:
Upstream primer, P2-F:5 '-CCACGCTCACCGTGGGGCTGCTGCTCAT-3 ',
Downstream primer, P2-R:5 '-GAGGGGGTGAGTGCTGTGTTCTC-3 ';
3. pcr amplification
25.0 �� L amplification system: 2 �� TaqPCRMasterMix(are established purchased from Beijing hundred Tyke company by primer of primer pair P1 and P2) 12.5 �� L, ddH2O9.5 �� L, P1-F0.4 �� L, P1-R0.6 �� L, P2-F0.6 �� L, P2-R0.4 �� L, DNA profiling 1.0 �� L;
Response procedures is: 94 DEG C of denaturation 4min; 94 DEG C of sex change 30s, 64 DEG C of annealing 30s, 72 DEG C extend 15s, 35 circulations; 72 DEG C extend 10min;
4. the detection of pcr amplification product and result judge
Get 10.0 �� LPCR amplification after products, agarose gel electrophoresis (electrophoretic voltage: the 120V of 1.5%; Electrophoresis time: 40min) put sample, after electrophoresis terminates, with gel imaging system to sepharose photograph detection, electrophoretogram is see Fig. 1; Stripe size according to occurring in electrophoretogram judges: when there is 322bp and 224bp fragment, be then dominant white plumage homozygote (II genotype); When the fragment of 322bp and 144bp occurs, then it is wild-type homozygote (ii genotype); When the fragment of 322bp, 224bp and 144bp occurs, then it it is dominant white plumage heterozygote (Ii genotype);
(4) according to genotype result of determination, dominant Bai Yu site of selecting and remain is the individuality of ii genotype, eliminates the individuality that dominant Bai Yu site is II and Ii genotype, obtains the male and female chicken that dominant Bai Yu site is ii genotype;
(5) the blue or green shin white plumage male and female chicken selfing that dominant Bai Yu site is ii genotype is got, the method of step (3) is utilized to measure the genotype of self progeny, repeat above-mentioned steps to genetic stability, obtain the blue or green shank and recessive white feather chicken strain that dominant Bai Yu site is ii genotype;
(6) the blue or green shank and recessive white feather chicken strain that dominant Bai Yu site step (5) obtained is ii genotype is supporting with blue or green shin yellow pockmarked feather chicken mixing breed as male parent, produces coloured plumage broiler chicken.
Claims (8)
1. the selection of a blue or green shank and recessive white feather chicken strain, it is characterised in that: comprise the following steps:
(1) utilizing blue or green shank and recessive white feather chicken as male parent, Bai Laihang chicken hybridizes as female parent, produces F1 generation;
(2) F1 generation selfing obtains F2 generation, the F2 generation blue or green white plumage male and female chicken of shin of selecting and remain;
(3) AS-PCR is utilized to measure the genotype of the F2 generation blue or green white plumage male and female chicken of shin, dominant Bai Yu site of selecting and remain is the individuality of ii genotype, eliminate the individuality that dominant Bai Yu site is II and Ii genotype, obtain the white plumage male and female chicken of blue or green shin that dominant Bai Yu site is ii genotype;
(4) the blue or green shin white plumage male and female chicken selfing that dominant Bai Yu site is ii genotype is got, the method of step (3) is utilized to measure the genotype of self progeny, repeat above-mentioned steps to genetic stability, obtain the blue or green shank and recessive white feather chicken strain that dominant Bai Yu site is ii genotype;
Described step (3) utilize AS-PCR measure the method for F2 generation blue or green shin white plumage male and female chicken genotype, comprise the following steps: taking comprise PMEL17 gene chicken complete genome DNA to be measured as template, increase chicken PMEL17 gene I allelotrope by primer PCR of primer pair P1, and increase chicken PMEL17 gene i allelotrope by primer PCR of primer pair P2; The fragment that amplification obtains is carried out agarose gel electrophoresis, identifies the genotype in the dominant Bai Yu site of chicken according to agarose gel electrophoresis result;
Described primer pair P1 is:
Upstream primer, P1-F:5 '-CCGGCCTCTACTGCCTCAACGTCTCGT-3 ',
Downstream primer, P1-R:5 '-GTGCCCAGAGCTTCGGCGATGAGCGGTG-3 ';
Described primer pair P2 is:
Upstream primer, P2-F:5 '-CCACGCTCACCGTGGGGCTGCTGCTCAT-3 ',
Downstream primer, P2-R:5 '-GAGGGGGTGAGTGCTGTGTTCTC-3 ';
The described method identifying the dominant white plumage loci gene type of chicken is: II genotype shows as 322bp and 224bp band; Ii genotype shows as 322bp, 224bp and 144bp band; Ii genotype shows as 322bp and 144bp band.
2. the selection of blue or green shank and recessive white feather chicken strain according to claim 1, it is characterised in that: in described step (1), blue or green shank and recessive white feather chicken is the blue or green shin white feather chicken after blue or green shin jute plumage indigenous chicken suddenlys change.
3. the selection of blue or green shank and recessive white feather chicken strain according to claim 1, it is characterised in that: described pcr amplification reaction system is: 2 �� TaqPCRMasterMix12.5 �� L, ddH2O9.5 �� L, P1-F0.4 �� L, P1-R0.6 �� L, P2-F0.6 �� L, P2-R0.4 �� L, DNA profiling 1.0 �� L.
4. the selection of blue or green shank and recessive white feather chicken strain according to claim 1, it is characterised in that: described pcr amplification reaction program is: 94 DEG C of denaturation 4min; 94 DEG C of sex change 30s, 64 DEG C of annealing 30s, 72 DEG C extend 15s, 35 circulations; 72 DEG C extend 10min.
5. the selection of blue or green shank and recessive white feather chicken strain according to claim 1, it is characterised in that: the mass concentration of described sepharose is 1.5%.
6. the selection of blue or green shank and recessive white feather chicken strain according to claim 1, it is characterised in that: the voltage of described agarose gel electrophoresis is 120V, and electrophoresis time is 40min.
7. the selection of blue or green shank and recessive white feather chicken strain according to claim 1, it is characterized in that: the blue or green shank and recessive white feather chicken strain that dominant Bai Yu site step (4) obtained is ii genotype is supporting with blue or green shin yellow pockmarked feather chicken mixing breed, produces coloured plumage broiler chicken.
8. the selection of blue or green shank and recessive white feather chicken strain according to claim 7, it is characterised in that: described blue or green shin yellow pockmarked feather chicken kind is gu-shi chicken, limit chicken, Huaibei partridge chicken, Huainan Spotted-brown chicken, Wuhua chicken, Chongren fiber crops chicken, Lang Yaji, Lushi chicken, Land of Peach Blossoms chicken, precious jade chicken or camellia chicken.
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| CN104293905B (en) * | 2014-04-11 | 2016-06-01 | 河南农业大学 | A kind of primer for detecting the blue or green chain SNP site genotype of shin proterties of chicken, test kit and detection method |
| CN104719253A (en) * | 2015-04-15 | 2015-06-24 | 广西南宁市富凤农牧有限公司 | Breeding method of low-speed type super-quality green-leg parental breeding chickens |
| CN104823919B (en) * | 2015-05-18 | 2017-03-15 | 贵州省畜牧兽医研究所 | The breeding method of southeast of Guizhou Province Fructus Foeniculi chicken recessive white feather new lines and application |
| CN108220408B (en) * | 2018-01-12 | 2021-07-06 | 江苏省家禽科学研究所 | Grain-saving green-shin recessive white feather broiler new strain breeding method |
| CN111357714A (en) * | 2020-04-24 | 2020-07-03 | 广东天农食品有限公司 | Method for breeding green-foot partridge chickens |
| CN112063725B (en) * | 2020-09-14 | 2021-07-20 | 华南农业大学 | Chicken shin length and shin diameter related AGO3 gene molecular marker and application |
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