Primer pair and kit used for carrying out early stage gender authentication of chicken, and application of primer pair and kit
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A kit and a technology of specific primer pairs, applied in the field of primer pairs for early sex identification of chickens, can solve the problems of easy misjudgment, cumbersome experimental process, high toxicity, etc.
Pending Publication Date: 2021-05-04
JIANGSU INST OF POULTRY SCI
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However, the sequences of the two homologous copies of the CHD gene in female individuals are relatively conservative, generally only 100-200bp apart. Using agarose electrophor
Method used
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Embodiment 1
[0030] Embodiment 1, establishment of method
[0031] Samples for testing: 10 adult camellia chickens of known sex, 5 of which are numbered 1-5 are roosters, and 5 of which are numbered 6-10 are hens.
[0032] 1. Take the blood of the test sample and extract the genomic DNA.
[0033] 2. Using the genomic DNA obtained in step 1 as template DNA, perform PCR amplification.
[0034] The primer pairs used in PCR amplification are as follows:
[0035] Forward primer (SEQ ID NO.1): 5'-TGTATTTTGGTTCTACAGGC-3';
[0036] Reverse primer (SEQ ID NO. 2): 5'-ATCATCATAACCATAAATCT-3'.
[0037] PCR amplification reaction system (50 μL): 2×PCR Mix (Nanjing Boerdi Biological Co., Ltd.) 25 μL, 10 μmol / L forward primer 1 μL, 10 μmol / L reverse primer 1 μL, 50-100 μg / ml template DNA 2 μL, Ultrapure water 21 μL.
[0038] The reaction program of PCR amplification: 95°C for 5min; 95°C for 30s, 60°C for 30s, 72°C for 30s, 35 cycles; 72°C for 10min.
[0039] 3. After step 2 is completed, the PCR am...
Embodiment 2
[0042] Embodiment 2, the application of method
[0043] Samples for testing: 17 Tibetan chicken chicks of unknown sex.
[0044] Method is with embodiment 1.
[0045] Electropherogram see figure 2 . figure 2 In , lanes 1 to 17 sequentially represent 17 samples for testing. Among them, some of the test samples showed a single characteristic band of 538bp (swimming lanes 1, 3, 6, 7, 8, 10, 11, 15, 17), which were judged to be roosters, and some of the test samples showed two characteristic bands of 960bp and 538bp (Swimming lanes 2, 4, 5, 9, 12, 13, 14, 16), judged to be a hen.
[0046] The test specimens were dissected and sexual organs were observed, and the above identification results were completely correct.
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Abstract
The invention discloses a primer pair and a kit used for carrying out early stage gender authentication of a chicken, and application of the primer pair and the kit. The invention provides a specific primer pair which consists of a single stranded DNA molecule disclosed by SEQ ID NO.1 and a single stranded DNA molecule disclosed by SEQ ID NO.2. The invention also provides a method for authenticating the gender of the chicken. The method comprises the following steps of: taking the genome DNA of a chicken to be detected as a template, adopting the specific primer pair to carry out PCR amplification, if an amplification product is obtained, proving that the chicken to be detected is a cock or is a candidate cock, and if two amplification products are obtained, proving that the chicken to be detected is a hen or is a candidate hen. Compared with the prior art, the invention has the following advantages that the hen and the cock have a difference of band amounts, and the difference of characteristic bands is 400bp. Through agarose electrophoresis detection, operation is simple and convenient, wrong judgment is unlikely to occur, an authentication result is quick and accurate, and therefore, the primer pair, the kit and the method can be conveniently popularized and used by the grass roots and have a wide market application prospect.
Description
technical field [0001] The invention belongs to the field of biotechnology, and relates to a primer pair, a kit and applications thereof for early sex identification of chickens. Background technique [0002] Most commercial poultry breeds are produced in the form of matching lines. The male line generally grows faster, and the female line generally has a higher reproductive efficiency. Therefore, early sex identification is required. The male line must eliminate female chicks in advance, and the female line must eliminate male chicks in advance. Commercial broiler roosters grow faster, and males and females are reared in groups, and the breeding benefits will be better. When the commercial substitute layer hens are hatched, only the female chicks are kept, and the male chicks are directly eliminated. Therefore, early sex identification of chickens is of great significance in production. [0003] At present, in addition to methods such as sex-linked inheritance (gold and s...
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