CN106701935A - Fast pyramiding breeding method of high-quality broiler multi-target traits - Google Patents

Fast pyramiding breeding method of high-quality broiler multi-target traits Download PDF

Info

Publication number
CN106701935A
CN106701935A CN201611183590.1A CN201611183590A CN106701935A CN 106701935 A CN106701935 A CN 106701935A CN 201611183590 A CN201611183590 A CN 201611183590A CN 106701935 A CN106701935 A CN 106701935A
Authority
CN
China
Prior art keywords
receptor
primer sequence
upstream
dopamine receptor
chicken
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
CN201611183590.1A
Other languages
Chinese (zh)
Other versions
CN106701935B (en
Inventor
赵振华
黎寿丰
李春苗
黄华云
王钱保
吴兆林
刘帆
刘一帆
赵东伟
梁忠
卢建
李新
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Jiangsu Institute Poultry Sciences
Original Assignee
Jiangsu Institute Poultry Sciences
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Jiangsu Institute Poultry Sciences filed Critical Jiangsu Institute Poultry Sciences
Priority to CN201611183590.1A priority Critical patent/CN106701935B/en
Publication of CN106701935A publication Critical patent/CN106701935A/en
Application granted granted Critical
Publication of CN106701935B publication Critical patent/CN106701935B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
    • C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/124—Animal traits, i.e. production traits, including athletic performance or the like
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q2600/00—Oligonucleotides characterized by their use
    • C12Q2600/156—Polymorphic or mutational markers

Landscapes

  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Organic Chemistry (AREA)
  • Zoology (AREA)
  • Wood Science & Technology (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Microbiology (AREA)
  • Immunology (AREA)
  • Molecular Biology (AREA)
  • Biotechnology (AREA)
  • Biophysics (AREA)
  • Physics & Mathematics (AREA)
  • Biochemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

The invention discloses a fast pyramiding breeding method of high-quality broiler multi-target traits. The fast pyramiding breeding method comprises the following steps: 1, extracting of genome DNA of broiler; 2, primer designing: designing a polymerase chain reaction amplification primer according to gene sequences of a growth differentiation factor 9, an estrogen receptor, a dopamine receptor 1, a dopamine receptor 2 and a vasoactive intestinal peptide receptor 2; 3, polymerase chain reaction; 4, single-nucleotide polymorphism reaction; 5, SSCP analysis on a target fragment, development by use of silver staining after gel electrophoresis to obtain an SSCP map of the DNA, selection of the growth differentiation factor 9, the estrogen receptor, the dopamine receptor 1, the dopamine receptor 2, the vasoactive intestinal peptide receptor 2 and individuals with genotypes of genes of QQ, GG, KK, TT and TT according to the banding pattern selection in 5 maps. The fast pyramiding breeding method disclosed by the invention is obvious in effect, the detection method is simple and fast and cannot be influenced by external environments, and an efficient and fast high-quality broiler pyramiding breeding new method is formed.

Description

A kind of rapid polymerization breeding method of quality broiler chicken multiple target character
Technical field
The present invention relates to quality broiler chicken multiple target character multiple gene polymerization breeding method, belong to poultry breeding method, specifically Be related to the assembling of multiple target character and be polymerized, the choosing such as quick fixed and homozygosis of multiple gene polymerization character screening, genes of interest type Educate method.
Background technology
Quality broiler chicken animal husbandry is the specialty industries of China, and the eternal target of quality broiler chicken breeding is " efficiently and high-quality ". Compared with the large-scale white meat-type chickens of state's extra income, China's quality broiler chicken will not only have in the proterties such as flavor substance content and meat quality High requirement, but also require the characteristics of breeder has reproductive performance higher, and commercial generation chicken has the appropriate speed of growth.Breeder energy More commercial generation seedling chicken numbers and the appropriate speed of growth of commercial generation chicken be provided and excellent meat be quality broiler chicken " efficiently with it is excellent The specific manifestation of matter " property.Although recent years take with the breeder more former Local chicken breeds of reproductive trait that egg production is representative index Obtained larger Breeding Progress, but qualified hatching egg number, chickling-surviving rate etc. represent hatching efficiency proterties it is still relatively low, as the production of hybrid seeds is looked forward to The problem sooner or later day of the aggravation competed between industry, the breeding cost of quality broiler chicken and commercial generation chicken listing age in days is aobvious important.How Selection raising is carried out to influence " efficient " major traits with classical quantitative inheritance theory and Modern Molecular Biotechnology, is sought The equalization point of " high-quality " and " efficient " is looked for, is one of key technology that quality broiler chicken breeding field needs innovation and breakthrough.
Sexal maturity is to determine one of fundamental prerequisite that quality broiler chicken is listed.Such as to reach certain sexal maturity at present will Ask, the feeding period of commercial generation chicken is then relatively long.Research in terms of to chicken sexal maturity proterties both at home and abroad mainly has temperature, latitude The climatic factors such as degree, season, illumination, feed nutrition etc. raise factor, and body weight, levels of reproductive hormones and inherent cause are to sexal maturity Influence, the genetic parameter estimation of influence and sexal maturity correlated traits of the sexal maturity to meat etc..It is main in terms of molecular studies Concentrate on QTL positioning and single candidate dna variation with the correlation of sex premature proterties on.It was found that relevant with Age at first laying QTL be located at chicken No. l, No. 3 and Z chromosome.As sex premature Candidate Gene Study it is earliest dw genes, is followed by chicken Borne virus gene.In recent years, gradually it is applied to candidate gene approach in the economic characters research of chicken, it is increasing to wait Gene is selected to be found relevant with the sex premature proterties of chicken.Existing research is it has been shown that the size of cockscomb can be effectively reactive Maturation time.
China's requirement mostly important to quality broiler chicken be：To be preced with that big, whisker is red, feather is bright during listing.This is to be different from The characteristics of one important mark of the large-scale broiler chicken of state's extra income is also compatriots' consumption demand.The sex premature of chicken can have influence on chicken The secondary sex characters such as size, the feather color and luster of hat；Meanwhile, there is research to confirm, when sexal maturity, body composition changes chicken Become, carcass quality is improved, the flavor substance content such as intramuscular fat is improved, these flavor substances are that muscle taste is aromatic, succulence With the material base of local flavor, the chicken of early sexual maturity, chicken flavor is better.The large-scale meat quality of table poultry of state's extra income it is not good also exactly because For its growth period is short, the slow reason of sexal maturity.
Just country's general status is seen at present, and the father and mother with suitable market needs are for chicken Gao Fan, commercial generation chicken precocity, raising Phase is shorter, and the quality broiler chicken new varieties (breed system) that can combine " high-quality with efficiently " feature are also few, it is impossible to meet China excellent The need for matter Chicken industry is produced at this stage.The need for the broiler chicken of the precocious numerous high-quality high of cultivation is whole industry development, but often New varieties (being) cycle that rule breeding is cultivated is more long, general more than 5 years, and how huge expends, it is desirable to which the index of measure is more, flower Take artificial many.The present invention overcomes conventional method cycle shortcoming long, the excellent new varieties of quick initiative Comprehensive Traits in 1-2 (being), greatly speeds up the efficiency and process of breeding.
The content of the invention
It is an object of the invention to provide a kind of rapid polymerization breeding method of multiple target character, the inventive method effect shows Write, detection method simple and fast, and it is free from the influence of the external environment.This technology is overcome in quality broiler chicken conventional selection breeding The problem that genotype homozygosis is slow, character determination efficiency is low, gives full play to effect of the biotechnology in poultry breeding, so as to be formed A kind of efficient, quick quality broiler chicken pyramiding breeding new technology.
The purpose of the present invention is achieved through the following technical solutions, a kind of rapid polymerization breeding side of multiple target character Method, comprises the following steps：
1st, the extracting genome DNA of chicken；
2nd, design of primers：According to growth differentiation factor 9 (GDF9), ERs (ESR), Dopamine receptor_1 (DRA1), Dopamine receptor 2 (DRA2), vip receptor 2 (VIP2) gene order design PCR (PCR) amplification Primer；
Growth differentiation factor 9 (GDF9)-tggagggtacgtggaggat-3 ' of upstream primer sequence 5 ', downstream primer sequence 5’-cacacctacctgtaatcggt-3’；
ERs (ESR) upstream primer sequence：5 '-gatgcacgcatagcttttca-3 ', downstream primer sequence： 5’-acggaaaagagtcagcctca-3’；
Dopamine receptor_1 (DRA1) upstream primer sequence：5 '-gctgtggctgagtttccttc-3 ', anti-sense primer sequence Row：5’-acccttggaacacactgagg-3’；
Dopamine receptor 2 (DRA2) upstream primer sequence：5’-aacccaccctcctcagactt-3’；Anti-sense primer sequence Row：5’-ggatggagatgggagacaga-3’；
Vip receptor 2 (VIP2) gene order upstream primer sequence：5’-gttggggagtggaggttcag- 3’；Downstream primer sequence：5’-attgggctggagatggcaaa-3’；
3rd, PCR (PCR)；Using 20 μ L reaction systems：10 × buffer is added in PCR reaction tubes (25mmol/L)2μL；Mg2+(10pmol/μL)2.2μL；The μ L of dNTPs (2.5mmol/L) 0.8, DNA profiling 1 μ L, it is any of the above-described To each 1 μ L of upstream and downstream primer (being 10pmol/L) of primer, Taq DNA polymerase (5U/ μ L) 0.2 μ L, plus ultra-pure water is to 20 μ L, of short duration centrifugation after fully mixing；PCR reaction tubes are put into PCR instrument carries out reaction amplification：94 DEG C of predegeneration 5min；94 DEG C of denaturation 30s, anneal 30s, 72 DEG C of extension 30s, 30 circulations；Last 72 DEG C of extensions 10min, 4 DEG C of preservations, 5 pairs of annealing temperatures of primer Respectively 58 DEG C, 60 DEG C, 60 DEG C, 59 DEG C and 61 DEG C.
4th, SNP reaction (SSCP)；2 μ L pcr amplification products add 7 μ L sample-loading buffers, 98 DEG C of denaturation 10min, then ice bath 5min.5 pairs of polyacrylamide concentrations of primer are 10%, and gel cross-linkage degree is 29：1, voltage is all It is 150V, electrophoresed overnight；
5. purpose fragment is developed the color after gel electrophoresis through sscp analysis with silver staining, obtains the SSCP collection of illustrative plates of DNA, according to 5 figures Banding pattern in spectrum selects growth selection differentiation factor 9 (GDF9), ERs (ESR), Dopamine receptor_1 (DRA1), DOPA Amine receptor 2 (DRA2), the genotype of vip receptor 2 (VIP2) gene are respectively QQ, GG, KK, TT, TT genotype It is individual；
6. each gene pleiomorphism and cockscomb, reproductive trait correlation analysis.It is determined that polymerization genotype is the individual of QQGGKKTTTT Body comb growth is most fast, and egg number is maximum, is reserved seed for planting in selection QQGGKKTTTT genotype individuals；
7. select and remain the male and female chicken apolegamy of QQGGKKTTTT genotype individuals, genotype fixes, and its offspring will not produce gene Segregation phenomenon, make the reproductive performance of offspring on 5 genotype effects have optimum efficiency, and stabilization heredity, make cockscomb and Reproductive performance gene can reach homozygosis within 1 generation, accelerate the cultivation of the precocious and high Breeding lines of quality broiler chicken.
Compared with prior art, the invention has the advantages that：Overcome genotype homozygosis in conventional selection breeding Slowly, the low problem of character determination efficiency a, generation just completes gene pyramiding, and multiple gene polymerization breeding method is in 10 week old cockscombs Area, 43 weeks egg numbers and 66 weeks have been respectively increased 22.49mm on egg number2, 2.6 and 2.9.And multiple gene polymerization side Method is simple to operate, be able to can just be chosen seeds when shell is gone out, and reduces the quantity of population measure, has saved cost and manpower.
Brief description of the drawings
Fig. 1 is the SSCP collection of illustrative plates of GDF9 genes.In figure：1：QQ types；2：PP types；3：PQ types；4,5：OQ types；6,8：OP types； 7：OO types.
Fig. 2 is the sequencer map of GDF9 genes.
Fig. 3 is the SSCP collection of illustrative plates of ESR genes.In figure：1,2：GG types；5,6：AA types；3,4：AG types
Fig. 4 is the sequencer map of ESR genes.
Fig. 5 is the SSCP collection of illustrative plates of DRA1 genes.In figure：1,2,6：KK；3,5：LL；4：KL
Fig. 6 is the sequencer map of DRA1 genes.
Fig. 7 is the SSCP collection of illustrative plates of DRA2 genes.In figure：2,3,4,6,7,9：CC types；5,10：TT types；1,11：CT types
Fig. 8 is the sequencer map of DRA2 genes.
Fig. 9 is the SSCP collection of illustrative plates of DRA2 genes.In figure：2,5,7,8：CC types；4：TT types；1,3,6：CT types
Figure 10 is the sequencer map of DRA2 genes.
Figure 11 is the schematic diagram of quality broiler chicken multiple target character rapid polymerization breeding method.
Specific embodiment
As shown in figure 11, a kind of rapid polymerization breeding method of multiple target character, comprises the following steps：
1st, the extracting genome DNA of chicken；
2nd, design of primers：According to growth differentiation factor 9 (GDF9), ERs (ESR), Dopamine receptor_1 (DRA1), Dopamine receptor 2 (DRA2), vip receptor 2 (VIP2) gene order design PCR (PCR) amplification Primer；
Growth differentiation factor 9 (GDF9)-tggagggtacgtggaggat-3 ' of upstream primer sequence 5 ', downstream primer sequence 5’-cacacctacctgtaatcggt-3’；
ERs (ESR) upstream primer sequence：5 '-gatgcacgcatagcttttca-3 ', downstream primer sequence： 5’-acggaaaagagtcagcctca-3’；
Dopamine receptor_1 (DRA1) upstream primer sequence：5 '-gctgtggctgagtttccttc-3 ', anti-sense primer sequence Row：5’-acccttggaacacactgagg-3’；
Dopamine receptor 2 (DRA2) upstream primer sequence：5’-aacccaccctcctcagactt-3’；Anti-sense primer sequence Row：5’-ggatggagatgggagacaga-3’；
Vip receptor 2 (VIP2) gene order upstream primer sequence：5’-gttggggagtggaggttcag- 3’；Downstream primer sequence：5’-attgggctggagatggcaaa-3’；
3rd, PCR (PCR)；Using 20 μ L reaction systems：Upstream and downstream primer is added to mix in PCR reaction tubes The μ L of compound 2.0, the μ L of 10 × buffer (25mmol/L) 2；Mg2+(10pmol/μL)2.2μL；The μ L of dNTPs (2.5mmol/L) 0.8, DNA profiling 1 μ L, any of the above-described each 1 μ L of upstream and downstream primer (being 10pmol/L) to primer, Taq DNA polymerase (5U/ μ L) 0.2 μ L, plus ultra-pure water is to 20 μ L, of short duration centrifugation after fully mixing；PCR reaction tubes are put into PCR instrument carries out reaction expansion respectively Increase：：94 DEG C of predegeneration 5min；94 DEG C of denaturation 30s, anneal 30s, 72 DEG C of extension 30s, 30 circulations；Last 72 DEG C of extensions 10min, 4 DEG C of preservations, the annealing temperature of 5 pairs of primers is respectively 58,60,60,59 and 61 DEG C.
4th, SNP reaction (SSCP)；2 μ L pcr amplification products add 7 μ L sample-loading buffers, 98 DEG C of denaturation 10min, then ice bath 5min.5 pairs of polyacrylamide concentrations of primer are 10%, and gel cross-linkage degree is 29：1, voltage is all It is 150V, electrophoresed overnight；
5. purpose fragment is through sscp analysis, is developed the color with silver staining after gel electrophoresis, obtain DNA SSCP collection of illustrative plates (see Fig. 1- 10), the banding pattern in 5 collection of illustrative plates selects growth selection differentiation factor 9 (GDF9), ERs (ESR), dopamine receptor 1 (DRA1), dopamine receptor 2 (DRA2), the genotype of vip receptor 2 (VIP2) gene be respectively QQ, GG, KK, The individuality of TT, TT genotype；
6. each gene pleiomorphism and cockscomb, reproductive trait correlation analysis.It is determined that polymerization genotype is the individual of QQGGKKTTTT Body comb growth is most fast, and egg number is maximum, is reserved seed for planting in selection QQGGKKTTTT genotype individuals；
7. select and remain the male and female chicken apolegamy of QQGGKKTTTT genotype individuals, genotype fixes, and its offspring will not produce gene Segregation phenomenon, make the reproductive performance of offspring on 5 genotype effects have optimum efficiency, and stabilization heredity, make cockscomb and Reproductive performance gene can reach homozygosis within 1 generation, accelerate the cultivation of the precocious and high Breeding lines of quality broiler chicken.
Embodiment：Quality broiler chicken is precocious, numerous S3 systems high seed selection.
Proterties with cockscomb size and the uniformity, 43 weeks egg numbers and 66 weeks egg numbers as seed selection.
Conventional selection：Using individual choice, the system of selection of families selecting.Hen is selected and remain rate 25~35%, and cock is selected and remain Rate 5~7%.Each generation locking is bred, it is to avoid 1/4 inbred.
Seed selection program：
1. wing number is worn by pedigree when going out shell；
The full group's measurement cockscomb size of 2.10 week old, the average weight and coefficient of variation size of per stirpes are ranked up, eliminate The family that cockscomb average value is too small and the coefficient of variation is too big；
3. in the family selected and remain, all individualities are ranked up by cockscomb size, eliminate 10% body weight cockscomb maximum Individuality, cockscomb lower limit is then determined according to the rate of selecting and remain, carry out selection and stay.
4th, hen carries out individuality and lays eggs record to 66 weeks.
5th, according to 43 weeks and the egg production of 66 weeks, family of selecting and remain eliminates the few family of egg production, in the family selected and remain In, all individualities are ranked up by egg number size, determine that hen is laid eggs lower limit, is carried out selection and is stayed according to the rate of selecting and remain.
6th, new family, successive propagation are set up by non-close relative after 43 weeks.
Multiple gene polymerization breeding selection：Multiple gene polymerization method for selecting molecular marker
1. wing number is worn by pedigree when going out shell；Wing venous are taken a blood sample
2. STb gene is extracted, is carried out by inventive step method.
The S3 systems conventional herd breeding Breeding Progress of table 1
The S3 systems multiple gene polymerization Breeding Progress of table 2
From table 1-2, conventional herd breeding method is distinguished on 10 week old cockscomb areas, 43 weeks egg numbers and 66 weeks egg numbers Improve 9.21mm2, 2.2 and 2.3, and multiple gene polymerization breeding method 10 week old cockscomb areas, 43 weeks egg numbers and 22.49mm has been respectively increased on 66 weeks egg numbers2, 2.6 and 2.9.Although the seed selection result difference between 2 kinds of methods does not show Write, but multiple gene polymerization method is slightly better than Conventional methods of selection, and also multiple gene polymerization method is simple to operate, and can be when shell be gone out Can just choose seeds, reduce the quantity of raising, save cost and manpower.
A kind of rapid polymerization breeding method of quality broiler chicken multiple target character
<110>Jiangsu Inst. of Fowls Science
<120>A kind of rapid polymerization breeding method of quality broiler chicken multiple target character
<160> 10
<210> 1
<211> 19
<212> DNA
<213>Chicken
<220>
<221>Growth differentiation factor 9 sense primer
<400> 1
tggagggtac gtggaggat 19
<210> 2
<211> 20
<212> DNA
<213>Chicken
<220>
<221>Growth differentiation factor 9 anti-sense primer
<400> 2
cacacctacc tgtaatcggt 20
<210> 3
<211> 20
<212> DNA
<213>Chicken
<220>
<221>ERs sense primer
<400> 3
gatgcacgca tagcttttca 20
<210> 4
<211> 20
<212> DNA
<213>Chicken
<220>
<221>ERs anti-sense primer
<400> 4
acggaaaaga gtcagcctca 20
<210> 5
<211> 20
<212> DNA
<213>Chicken
<220>
<221>Dopamine receptor_1 sense primer
<400> 5
gctgtggctg agtttccttc 20
<210> 6
<211> 20
<212> DNA
<213>Chicken
<220>
<221>Dopamine receptor_1 anti-sense primer
<400> 6
acccttggaa cacactgagg 20
<210> 7
<211> 20
<212> DNA
<213>Chicken
<220>
<221>The sense primer of dopamine receptor 2
<400> 7
aacccaccct cctcagactt 20
<210> 8
<211> 20
<212> DNA
<213>Chicken
<220>
<221>The anti-sense primer of dopamine receptor 2
<400> 8
ggatggagat gggagacaga 20
<210> 9
<211> 20
<212> DNA
<213>Chicken
<220>
<221>The gene order sense primer of vip receptor 2
<400> 9
gttggggagt ggaggttcag 20
<210> 10
<211> 20
<212> DNA
<213>Chicken
<220>
<221>The gene order sense primer of vip receptor 2
<400> 10
attgggctgg agatggcaaa 20

Claims (3)

1. a kind of rapid polymerization breeding method of quality broiler chicken multiple target character, it is characterized in that, comprise the following steps：
（1）Extract the genomic DNA of chicken；
（2）Design of primers：Growth differentiation factor 9, ERs, Dopamine receptor_1 according to chicken, dopamine receptor 2, blood vessel Active intestines peptide receptor 2 gene sequences Design PCR amplification primer；
- the tggagggtacgtggaggat-3 ' of growth differentiation factor 9 upstream primer sequence 5 ', downstream primer sequence 5 '- cacacctacctgtaatcggt-3’；
ERs upstream primer sequence：5 '-gatgcacgcatagcttttca-3 ', downstream primer sequence：5’- acggaaaagagtcagcctca-3’
Dopamine receptor_1 upstream primer sequence：5 '-gctgtggctgagtttccttc-3 ', downstream primer sequence：5’- acccttggaacacactgagg-3’；
The upstream primer sequence of dopamine receptor 2：5’-aacccaccctcctcagactt-3’；Downstream primer sequence：5’- ggatggagatgggagacaga-3’；
The gene order upstream primer sequence of vip receptor 2：5’-gttggggagtggaggttcag-3’；Anti-sense primer Sequence：5’-attgggctggagatggcaaa -3’；
（3）PCR；Using 20 μ L reaction systems：The μ of 25 10 × buffer of mmol/L 2 in PCR reaction tubes L；10 pmol/μL Mg2+2.2 μL；μ L, 10 the pmol/L growths point of 2.5 mmol/L dNTPs, 0.8 μ L, DNA templates 1 Change each 1 μ L of the upstream and downstream primer of the factor 9 or 10 each 1 μ L or 10 of pmol/L ERs upstream and downstream primer Each 1 μ L of pmol/L Dopamine receptor_1s upstream and downstream primer or each 1 μ L of the upstream and downstream primer of 10 pmol/L dopamine receptors 2, Or each 1 μ L of 10 pmol/L vip receptors, 2 gene order upstream and downstream primer, 5 U/ μ L TaqDNA polymerases 0.2 μ L, plus ultra-pure water, to 20 μ L, PCR reaction tubes are put into PCR instrument carries out reaction amplification respectively；
（4）SNP reacts；2 μ L PCR amplified productions add 7 μ L sample-loading buffers, 98 DEG C of denaturation 10 Min, the then min of ice bath 5；Overnight gel electrophoresis；
（5）Purpose fragment is developed the color after gel electrophoresis through sscp analysis with silver staining, the SSCP collection of illustrative plates of DNA is obtained, according to 5 collection of illustrative plates In banding pattern selection growth selection differentiation factor 9, ERs, Dopamine receptor_1, dopamine receptor 2, vasoactive intestinal peptide The genotype of receptor 2 gene is respectively the individuality of QQ, GG, KK, TT, TT genotype.
2. the rapid polymerization breeding method of quality broiler chicken multiple target character according to claim 1, it is characterized in that, it is described Pcr amplification reaction program is：94 DEG C of min of predegeneration 5；94 DEG C of 30 s of denaturation, anneal 30 s, 72 DEG C of 30 s of extension, 30 circulations；10 min of last 72 DEG C of extensions, 4 DEG C of preservations, the annealing temperature of 5 pairs of primers be respectively 58 DEG C, 60 DEG C, 60 DEG C, 59 DEG C and 61 DEG C.
3. the rapid polymerization breeding method of quality broiler chicken multiple target character according to claim 1, it is characterized in that, step （4）Described in gel electrophoresis 5 pairs of polyacrylamide concentrations of primer be 10 %, gel cross-linkage degree is 29：1, voltage All it is 150V.
CN201611183590.1A 2016-12-20 2016-12-20 Rapid multi-target character polymerization breeding method for high-quality broiler chickens Active CN106701935B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201611183590.1A CN106701935B (en) 2016-12-20 2016-12-20 Rapid multi-target character polymerization breeding method for high-quality broiler chickens

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201611183590.1A CN106701935B (en) 2016-12-20 2016-12-20 Rapid multi-target character polymerization breeding method for high-quality broiler chickens

Publications (2)

Publication Number Publication Date
CN106701935A true CN106701935A (en) 2017-05-24
CN106701935B CN106701935B (en) 2020-01-07

Family

ID=58938118

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201611183590.1A Active CN106701935B (en) 2016-12-20 2016-12-20 Rapid multi-target character polymerization breeding method for high-quality broiler chickens

Country Status (1)

Country Link
CN (1) CN106701935B (en)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110651756A (en) * 2019-10-28 2020-01-07 北京市华都峪口禽业有限责任公司 Multi-character seed selection method for livestock and poultry

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103436630A (en) * 2013-09-23 2013-12-11 四川省畜牧科学研究院 High-quality broiler chicken inosinic acid and intramuscular fat content related gene polymerization breeding method
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

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103436630A (en) * 2013-09-23 2013-12-11 四川省畜牧科学研究院 High-quality broiler chicken inosinic acid and intramuscular fat content related gene polymerization breeding method
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

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
H.Y. HUANG等: "Polymorphism identification in BMP15 and GDF9 genes and their association with egg production in chickens", 《BRITISH POULTRY SCIENCE》 *
张增荣等: "优质肉鸡肌苷酸与肌内脂肪含量相关基因聚合效应研究", 《四川农业大学学报》 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110651756A (en) * 2019-10-28 2020-01-07 北京市华都峪口禽业有限责任公司 Multi-character seed selection method for livestock and poultry
CN110651756B (en) * 2019-10-28 2021-08-10 北京市华都峪口禽业有限责任公司 Multi-character seed selection method for livestock and poultry

Also Published As

Publication number Publication date
CN106701935B (en) 2020-01-07

Similar Documents

Publication Publication Date Title
CN107494410B (en) Method for producing dark and light shell green-shell laying hens in matched combination manner
CN104823919B (en) The breeding method of southeast of Guizhou Province Fructus Foeniculi chicken recessive white feather new lines and application
CN106520939B (en) A kind of method and its application of madai Germplasm Identification
CN106755447B (en) Molecular marker-assisted selection method for reducing abdomen fat rate of white-feather king pigeon and young pigeon
CN105002267A (en) Penaeus japonicus molecule marking method and application
CN109182534A (en) Jian carp breeding of new variety method
CN103798169A (en) Method for quickly establishing improved breeding system of YY super-male pelteobagrus fulvidraco
CN101921859B (en) Method for breeding lean-type Chinese Huai pigs in multi-gene pyramiding manner based on growth traits thereof
CN107494415B (en) Method for breeding new Guangdong yellow chickens
CN105624318A (en) SNP site related to growth trait of paralichthys olivaceus as well as screening method and application of SNP site
CN113430277A (en) Primer group for identifying sex-linked dwarf gene and application thereof
CN104894289A (en) Kit for detecting chicken growth characters and chicken molecular breeding method
CN110551823B (en) SNP molecular marker related to sperm storage capacity of hen and application thereof
CN115251004B (en) Cultivation method of green-shank yellow-feather high-quality broilers
CN106701935A (en) Fast pyramiding breeding method of high-quality broiler multi-target traits
CN103966316B (en) A kind of diploid loach micro-satellite Parentage determination method and application thereof
CN111213613A (en) Molecular breeding method of black-feather green-foot powder-shell meat-egg dual-purpose chicken
CN114214432A (en) Molecular marker for identifying melanin diffusion gene locus of chicken and application of molecular marker in assisted breeding
CN109439771A (en) A method of hybridization porgy family is identified using microsatellite marker
CN110878363B (en) Detection method and application of correlation between VIPR1 gene and chicken testicular character
CN103416351A (en) Method for establishing Bian chicken quick-grow type AA strain and slow-grow type GG strain colony
CN110551824B (en) SNP molecular marker related to sperm storage capacity of hen and application thereof
CN111418548A (en) Breeding method of dwarf yellow chicken strain with homozygous mutation caused by pathogen
CN109943639B (en) SNP molecular marker for selecting hen semen storage capacity and application thereof
CN114208766B (en) Breeding method of non-dwarf Ningdu yellow chicken mating line

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
CB02 Change of applicant information
CB02 Change of applicant information

Address after: 225261 Jiangsu Province, Yangzhou city Jiangdu District Shaobo Town Street No. 2

Applicant after: Jinagsu Poultry lnstitute

Address before: 225125 Jiangsu province Hanjiang District of Yangzhou city CHANGJEI Road No. 58

Applicant before: Jinagsu Poultry lnstitute

CB02 Change of applicant information
CB02 Change of applicant information

Address after: 225125 No. 58, Cang Jie Road, Hanjiang District, Yangzhou, Jiangsu

Applicant after: Jinagsu Poultry lnstitute

Address before: 225261 2 street, Shaw Town, Jiangdu District, Yangzhou, Jiangsu

Applicant before: Jinagsu Poultry lnstitute

GR01 Patent grant
GR01 Patent grant