AU2019101693A4 - A Primer for Identification of Shaobo Chicken and Its Application and A Method for Rapid Identification of Shaobo Chicken - Google Patents
A Primer for Identification of Shaobo Chicken and Its Application and A Method for Rapid Identification of Shaobo Chicken Download PDFInfo
- Publication number
- AU2019101693A4 AU2019101693A4 AU2019101693A AU2019101693A AU2019101693A4 AU 2019101693 A4 AU2019101693 A4 AU 2019101693A4 AU 2019101693 A AU2019101693 A AU 2019101693A AU 2019101693 A AU2019101693 A AU 2019101693A AU 2019101693 A4 AU2019101693 A4 AU 2019101693A4
- Authority
- AU
- Australia
- Prior art keywords
- primer
- chicken
- shaobo
- band
- pcr amplification
- 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.)
- Ceased
Links
Classifications
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New breeds of animals
- A01K67/02—Breeding vertebrates
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K67/00—Rearing or breeding animals, not otherwise provided for; New breeds of animals
- A01K67/027—New breeds of vertebrates
-
- 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
-
- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
- A01K2227/00—Animals characterised by species
- A01K2227/30—Bird
-
- 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
Abstract
A primer comprises a primer Fl and a primer F2, and the nucleotide sequence of primer Fl and primer F2 are shown in the sequence list. The present invention also provides an application of the primer as a molecular identification marker for Shaobo Chicken germplasm resources. Furthermore, the present invention also provides a method for rapid identification of Shaobo Chicken, characterized in that: with DNA genome extracted by blood of detected chicken as a template, PCR amplification is carried out by using the primer according to claim 1, PCR amplification product is detected by agarose gel electrophoresis, DW and/or dw gene is judged according to band: only a 417 bp single band amplified indicates chicken with DW gene alone; only a 217 bp single band amplified indicates chicken with dw gene alone; both 417 bp and 217 bp bands amplified indicate chicken with both DW and dw genes; only 417 bp band indicates parent-paternal line, only 217 bp band indicates parent-maternal line, and both 417 bp and 217 bp bands indicate commercial generation. The present invention utilizes amplification band of different sizes of sex-linked dwarf gene (dw) to identify Shaobo Chicken, which is not affected by environment and has lots of advantages, such as simple operation, rapid detection, accurate result, strong universality and low cost.
Description
The present invention relates to an identification of parent and commercial generation of Shaobo Chicken, and more particularly to a method for rapid identification of Shaobo Chicken by using PCR technology, which belongs to the technical field of molecular biology.
Background Technology
Shaobo Chicken complete set line was conducted by Jiangsu Institute of Poultry Science through introduction of dw gene into high-quality local chicken breeds in China to breed a line of high-quality dwarf green shank spotted feather broiler (S2 line) for the first time at home and abroad and with participation of complete set line. The complete set line that has been approved by the national breed (complete set line) review (N09XPZZZ No. 12) is the first national high-quality broiler complete set line with proprietary intellectual property rights. The technical achievement has been identified at international advanced level. Sex-linked dwarf gene (dw) is a growth hormone receptor (GHR) gene deficient type, and Dw gene is an sex-associated gene located on Z chromosome, between in site of liver necrosis gene and we site of wingless gene, and near silver (S) and golden (s) feather gene and slow-feathering (K) and fast-feathering (k) gene. Normal gene DW is dominant for dw, so only dwarf homozygotic cock and hen carrying dwarf gene show dwarf character, dw and DW are incomplete alleles, and dw gene has the characteristics of quality and quantitative trait locus. Dwarf chicken is the phenotype of chicken homozygotic sex-linked dwarf gene (dw), which have lots of advantages, such as dwarf, low consumption, low basal metabolism, high egg production rate, high feeding density, high feed compensation and cost saving.
Compared with the same type of chicken breed, Shaobo Chicken complete set line has 10% earlier age at first egg, 20% higher egg laying performance and more than 25% feed saving per chick, and Shaobo Chicken complete set line has commercial generations with green shank, spotted feather and excellent appearance; precocious puberty, good meat quality and strong adaptability, which can not only be kept in hilly areas, but also suitable for large-scale cage culture; high market i
2019101693 24 Dec 2019 acceptance, about 15% - 20% higher selling price, and more than 10% - 15% higher comprehensive benefit of feeding. Due to very extensive prospect of market application of Shaobo Chicken, pseudo-Shaobo Chicken appears in many places, which needs a simple and rapid method for identification.
PCR is a kind of molecular biology technology, which is widely used in medical and biological laboratories, such as judging availability of a certain genetic disease atlas in samples, diagnosis of infectious diseases, gene replication and paternity test.
Invention Summary
The purpose of the present invention is to provide a primer for identification of Shaobo chicken and its application and a method for rapid identification of Shaobo Chicken. The present invention utilizes amplification band of different sizes of sex-linked dwarf gene (dw) to identify Shaobo Chicken, which is not affected by environment and has lots of advantages, such as simple operation, rapid detection, accurate result, strong universality and low cost.
The present invention provides a primer, comprising a primer Fl and a primer F2,
An upstream primer F of the primer Fl: 5’-tcccagactacacttctattca-3’;
A downstream primer R of the primer Fl: 5’-cgggacagatcaaagacaatac-3’;
An upstream primer F of the primer F2: 5’ -acctccaaagaaatctgtcgag-3’;
A downstream primer R of the primer F2: 5’-tggccaaatcctgaagtcct-3’.
The present invention also provides an application of the primer as a molecular identification marker for Shaobo Chicken germplasm resources.
Furthermore, the present invention also provides a method for rapid identification of Shaobo Chicken, characterized in that: with DNA genome extracted from blood of detected chicken breed as a template, PCR amplification is carried out by using the primer according to claim 1, PCR amplification product is detected by agarose gel electrophoresis, DW and/or dw gene is judged according to band: only a 417 bp single band amplified indicates chicken with DW gene alone; only a 217 bp single band amplified indicates chicken with dw gene alone; both 417 bp and 217 bp bands amplified indicate chicken with both DW and dw genes; only 417 bp band indicates parent-paternal line, only 217 bp band indicates parent-maternal line, both 417 bp and 217 bp bands indicates commercial generation; and only 417 bp single band of or 217 bp single band is present in
2019101693 24 Dec 2019 the PCR amplification product, indicating pseudo-Shaobo commercial chicken.
A reaction system of the PCR amplification is 20 pL, and the reaction system comprises the following components: 10 x PCR Buffer 2.0 pL, Mg2+ 2.2 pL, dNTP Mixture 0.8 pL, Taq DNA polymerase (5 U/pL) 0.2 pL, upstream and downstream primers of the primers Fl and F2 1 pL each, cDNA template 1 pL, and ddH2O 9.8 pL.
Reaction conditions of the PCR amplification include: pre-denaturation at 95°C for 5 min; denaturation at 95°C for 30 s, annealing at 61.4°C for 30 s, extension at 72 °C for 30 s, and total 30 cycles; extension at 72°C for 8 min; and storage at 4°C.
The agarose gel electrophoresis detection is performed as follows: 5 pL of PCR amplification product is mixed with 1 pL of 6 x Loading Buffer for 1.5% agarose gel electrophoresis under 120 V for 20 min, and photographed with gel imaging system.
Compared with the existing technologies, the present invention has the following benefits:
First, the present invention extracts chicken genome DNA from test samples and carries out PCR amplification to judge Shaobo Chicken according to the characteristics of 417 bp band alone in parent-parental line, 217 bp band alone in parent-maternal line and both 417 bp and 217 bp bands in commercial generation. The present invention directly takes DNA genome extracted from blood of tested chicken as a template, and utilizes PCR technology to detect fragment size of dw gene. The present invention has lots of advantages, such as simple operation, rapid detection, accurate result, strong universality and low cost.
Second, the present invention discloses the application of dwarf gene (dw) as a molecular identification marker for parents and commercial generations of Shaobo Chicken. The present invention includes an upstream and downstream primer mixture and PCR reaction reagents for PCR amplification of dw gene. The PCR reaction reagents include 10 x buffer, dNTP and rTaq enzyme. The present invention includes an upstream and downstream primer mixture for PCR amplification of dw gene and reagents required for PCR. The method of the present invention has remarkable effect, simple and rapid detection, and is not affected by external environment. The present invention is applied to the breed identification of Shaobo Chicken, mainly for identification of ancestral, parent and commercial generations of Shaobo Chicken. The present invention is reasonable in design, convenient in use, reliable in results and low in cost.
2019101693 24 Dec 2019
Brief Description of the Drawings
FIG.1 is an amplification diagram of parent-parental line cock;
FIG2 is an amplification diagram of parent-maternal line hen;
FIG3 is an amplification diagram of commercial generation.
Detailed Description of the Presently Preferred Embodiments
A primer comprises a primer Fl and a primer F2,
An upstream primer F of the primer Fl: 5’-tcccagactacacttctattca-3’;
A downstream primer R of the primer Fl: 5’-cgggacagatcaaagacaatac-3’;
An upstream primer F of the primer F2: 5’ -acctccaaagaaatctgtcgag-3’;
A downstream primer R of the primer F2: 5’-tggccaaatcctgaagtcct-3’.
An application of the primer as a molecular identification marker for Shaobo Chicken germplasm resources.
A method for rapid identification of Shaobo Chicken, characterized in that: with DNA genome extracted from blood of detected chicken breed as a template, PCR amplification is carried out by using the primer according to claim 1, PCR amplification product is detected by agarose gel electrophoresis, DW and/or dw gene is judged according to band: only a 417 bp single band amplified indicates chicken with DW gene alone; only a 217 bp single band amplified indicates chicken with dw gene alone; both 417 bp and 217 bp bands amplified indicate chicken with both DW and dw genes; only 417 bp band indicates parent-paternal line, only 217 bp band indicates parent-maternal line, both 417 bp and 217 bp bands indicates commercial generation; and only 417 bp single band of or 217 bp single band is present in the PCR amplification product, indicating pseudo-Shaobo commercial chicken.
A reaction system of the PCR amplification is 20 pL, and the reaction system comprises the following components: 10 x PCR Buffer 2.0 pL, Mg2+ 2.2 pL, dNTP Mixture 0.8 pL, Taq DNA polymerase (5 U/pL) 0.2 pL, upstream and downstream primers of the primers Fl and F2 1 pL each, cDNA template 1 pL, and ddH2O 9.8 pL.
Reaction conditions of the PCR amplification include: pre-denaturation at 95°C for 5 min; denaturation at 95°C for 30 s, annealing at 61.4°C for 30 s, extension at 72 °C for 30 s, and total 30
2019101693 24 Dec 2019 cycles; extension at 72°C for 8 min; and storage at 4°C.
The agarose gel electrophoresis detection is performed as follows: 5 pL of the PCR amplification product is mixed with 1 pL of 6 x Loading Buffer for 1.5% agarose gel electrophoresis under 120 V for 20 min, and photographed with gel imaging system.
Embodiment 1
1. Test materials
Shaobo Chickens used in the experiment were provided by Jiangsu Institute of Poultry Science, including 10 dwarf parent-maternal cocks (Zdw Zdw), 10 heterozygous commercial cocks (ZDw Zdw), 10 normal parent-parental cock (ZDw ZDw), 10 dwarf maternal hens (Zdw W) and 10 normal parental hens (ZDw W). Blood samples were intravenously collected from wings of all chicken with heparin sodium for anticoagulation.
2. Operating steps
PCR amplification: a reaction system of the PCR amplification is 20 pL: 10 x PCR Buffer 2.0 pL, Mg2+ 2.2 pL, dNTP Mixture 0.8 pL, Taq DNA polymerase (5 U/pL) 0.2 pL, upstream and downstream primers of the primers Fl and F2 1 pL each, cDNA template 1 pL, and ddlLO 9.8 pL. Reaction conditions: pre-denaturation at 95°C for 5 min; denaturation at 95°C for 30 s, annealing at 61.4°C for 30 s, extension at 72 °C for 30 s, and total 30 cycles; extension at 72°C for 8 min; and storage at 4°C.
Agarose gel electrophoresis detection: 5 pL of the PCR amplification product is mixed with 1 pL of 6 x Loading Buffer for 1.5% agarose gel electrophoresis under 120 V for 20 min, and photographed with gel imaging system.
3. Results
The amplification diagram of dwarf parent-maternal cock is shown in FIG. 2, the amplification diagram of heterozygous commercial cock is shown in FIG. 3, the amplification diagram of parent-parental cock is shown in FIG. 1, the amplification diagram of dwarf maternal hen is shown in FIG. 2, and the amplification diagram of normal parental hen is shown in FIG. 1. It can be seen that the size of PCR amplification fragment can be used as a method for breed identification of Shaobo Chicken.
Claims (6)
1. A primer, characterized in that: the primer comprises of a primer Fl and a primer F2,
An upstream primer F of the primer Fl: 5’-tcccagactacacttctattca-3’;
Adownstream primer R of the primer Fl: 5’ -cgggacagatcaaagacaatac-3’;
An upstream primer F of the primer F2: 5’ -acctccaaagaaatctgtcgag-3’;
A downstream primer R of the primer F2: 5’-tggccaaatcctgaagtcct-3’.
2. An application of the primer according to claim 1 as a molecular identification marker for Shaobo Chicken germplasm resources.
3. A method for rapid identification of Shaobo Chicken, characterized in that: with DNA genome extracted by blood of detected chicken as a template, PCR amplification is carried out by using the primer according to claim 1, the PCR amplification product is detected by agarose gel electrophoresis, DW and/or dw gene is judged according to band: only a 417 bp single band amplified indicates chicken with DW gene alone; only a 217 bp single band amplified indicates chicken with dw gene alone; both 417 bp and 217 bp bands amplified indicate chicken with both DW and dw genes; only 417 bp band indicates parent-paternal line, only 217 bp band indicates parent-maternal line, both 417 bp and 217 bp bands indicate commercial generation; and only 417 bp single band of or 217 bp single band is present in the PCR amplification product, indicating pseudo-Shaobo commercial chicken.
4. The method for rapid identification of Shaobo Chicken according to claim 2, characterized in that: a reaction system of the PCR amplification is 20 pL, and the reaction system comprises of the following components: 10 x PCR Buffer 2.0 pL, Mg2+ 2.2 pL, dNTP Mixture 0.8 pL, Taq DNA polymerase (5 U/pL) 0.2 pL, upstream and downstream primers of the primers Fl and F2 1 pL each, cDNA template 1 pL, and ddH2O 9.8 pL.
5. The method for rapid identification of Shaobo Chicken according to claim 4, characterized in that: reaction conditions of the PCR amplification include: pre-denaturation at 95°C for 5 min; denaturation at 95°C for 30 s, annealing at 61.4°C for 30 s, extension at 72 °C for 30 s, and total 30 cycles; extension at 72°C for 8 min; and storage at 4°C.
2019101693 24 Dec 2019
6. The method for rapid identification of Shaobo chicken according to claim 3, characterized in that: the agarose gel electrophoresis detection is performed as follows: 5 pL of the PCR amplification product is mixed with 1 pL of 6 x Loading Buffer for 1.5% agarose gel electrophoresis under 120 V for 20 min, and photographed with gel imaging system.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| AU2019101693A AU2019101693A4 (en) | 2019-12-24 | 2019-12-24 | A Primer for Identification of Shaobo Chicken and Its Application and A Method for Rapid Identification of Shaobo Chicken |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| AU2019101693A AU2019101693A4 (en) | 2019-12-24 | 2019-12-24 | A Primer for Identification of Shaobo Chicken and Its Application and A Method for Rapid Identification of Shaobo Chicken |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| AU2019101693A4 true AU2019101693A4 (en) | 2020-02-06 |
Family
ID=69284281
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| AU2019101693A Ceased AU2019101693A4 (en) | 2019-12-24 | 2019-12-24 | A Primer for Identification of Shaobo Chicken and Its Application and A Method for Rapid Identification of Shaobo Chicken |
Country Status (1)
| Country | Link |
|---|---|
| AU (1) | AU2019101693A4 (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113265455A (en) * | 2021-05-19 | 2021-08-17 | 嘉应学院 | Rapid identification method and primer set for sex of American partridges |
| CN113430277A (en) * | 2021-07-30 | 2021-09-24 | 中国农业大学 | Primer group for identifying sex-linked dwarf gene and application thereof |
| CN114568384A (en) * | 2021-11-23 | 2022-06-03 | 江苏省家禽科学研究所 | Seed production method of early-maturing grain-saving stress-resistant broiler chicken complete set system |
-
2019
- 2019-12-24 AU AU2019101693A patent/AU2019101693A4/en not_active Ceased
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113265455A (en) * | 2021-05-19 | 2021-08-17 | 嘉应学院 | Rapid identification method and primer set for sex of American partridges |
| CN113430277A (en) * | 2021-07-30 | 2021-09-24 | 中国农业大学 | Primer group for identifying sex-linked dwarf gene and application thereof |
| CN114568384A (en) * | 2021-11-23 | 2022-06-03 | 江苏省家禽科学研究所 | Seed production method of early-maturing grain-saving stress-resistant broiler chicken complete set system |
| CN114568384B (en) * | 2021-11-23 | 2023-02-03 | 江苏省家禽科学研究所 | Seed production method of early-maturing grain-saving stress-resistant broiler chicken complete set system |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| AU2019101693A4 (en) | A Primer for Identification of Shaobo Chicken and Its Application and A Method for Rapid Identification of Shaobo Chicken | |
| CN108220408B (en) | Grain-saving green-shin recessive white feather broiler new strain breeding method | |
| US20090126033A1 (en) | Method of selective breeding based on ob genotype | |
| US20120012065A1 (en) | Systems and Methods for Improving Protein and Milk Production of Dairy Herds | |
| CN105506086B (en) | The relevant SNP marker of chicken fertilization duration character and its application | |
| JP2014531208A (en) | Gene copy number polymorphism testing method | |
| WO2023001212A1 (en) | Gene chip, molecular probe combination and kit for analyzing sheep milk production performance, and use | |
| WO2023001209A1 (en) | Gene chip, molecular probe combination, test kit and application for analyzing sheep fat tails | |
| Yang et al. | Effects in cattle of genetic variation within the IGF1R gene on the superovulation performance and pregnancy rates after embryo transfer | |
| CN113699246B (en) | SNP molecular marker affecting pig feed conversion efficiency character and application thereof | |
| CN114107516B (en) | SNP (single nucleotide polymorphism) marker for evaluating backfat thickness of pig and detection method thereof | |
| CN112921101A (en) | Molecular marker related to sheep remaining feed intake and application thereof | |
| CN110079613B (en) | Molecular marker of heat stress tolerance of Holstein cattle and detection method | |
| CN112280874B (en) | Copy number variation molecular marker influencing pig backfat thickness on pig chromosome 11 and application | |
| CN112760387B (en) | SNP molecular marker related to total nipple number of pigs and application | |
| Fortes et al. | Genetic markers in transcription factors of differentially expressed genes associated with post-partum anoestrus predict pregnancy outcome in an independent population of beef cattle | |
| CN107779517A (en) | A kind of molecular labeling for influenceing Duroc kind pig lean meat percentage character and its application | |
| CN113699247A (en) | SNP molecular marker on pig No. 1 chromosome and related to pig remaining feed intake and application thereof | |
| AU2011205160A1 (en) | Method for improving efficiencies in livestock production | |
| CN104593358A (en) | Primer for identifying Shaobai chicken as well as application and method for quickly identifying Shaobai chicken | |
| CN111206092A (en) | Disease screening primer probe combination and application thereof, kit and detection method | |
| CN112266968B (en) | Copy number variation molecular marker influencing weight age in days and daily gain on pig chromosome 13 and application | |
| CN113293219B (en) | Specific primer of molecular marker of Mongolian sheep fertility-related gene BMPR1B and application thereof | |
| CN112458183B (en) | Copy number variation molecular marker on pig No. 3 chromosome and related to daily gain and weight and age on market of pig and application thereof | |
| CN111850139B (en) | Molecular marker located on pig chromosome 12 and related to formation of pig monocrchidism and application |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| FGI | Letters patent sealed or granted (innovation patent) | ||
| MK22 | Patent ceased section 143a(d), or expired - non payment of renewal fee or expiry |