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Primer, kit and method for rapid identification of pigeon gender

A technology for the identification of pigeons and kits, which is used in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., and can solve the problems of poor specificity of primers and methods, difficult to distinguish, and prone to false negatives. , to achieve the effect that the amplification results are intuitive and clearly distinguishable, the extraction steps are omitted, and the amplification results are intuitive and distinguishable.

Pending Publication Date: 2019-06-21
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to solve technical problems such as the poor specificity of the above-mentioned primers and methods for identifying the sex of pigeons, it is difficult to distinguish and prone to false negatives, the invention provides a primer, a kit and a method for quickly identifying the sex of pigeons

Method used

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  • Primer, kit and method for rapid identification of pigeon gender
  • Primer, kit and method for rapid identification of pigeon gender
  • Primer, kit and method for rapid identification of pigeon gender

Examples

Experimental program
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Effect test

Embodiment 1

[0044] The extraction of embodiment 1 blood or feather marrow sample DNA

[0045] (1) Extraction method of feather marrow sample DNA in feather follicles: due to rich in protein components in pigeon feathers, it is very difficult to effectively amplify; the inventor explored a variety of feather sample lysates in the early stage of research, and passed a large Experimental verification, and finally determine the effective and feasible feather sample lysate. Press the newly picked feathers along the root to extract the feather marrow and put them in a 1.5ml EP tube, package and mark the pigeon number, add 300 microliters of lysate A, heat at 95°C for 10 minutes, then add 30 microliters of lysate B to complete the sample After extraction, the supernatant is the genomic DNA of the feather plume sample. Take 1.5 μl of the supernatant for PCR amplification, and store the rest at -20°C for future use. Samples of blood or feather samples are valid at room temperature or refrigerated...

Embodiment 2

[0047] Embodiment 2CHD gene amplification primer screening and specificity experiment

[0048] In the present invention, the CHD gene is screened as the target gene for PCR amplification detection, and a plurality of sets of amplification primers are designed. The following takes the amplified fragment sizes of 450bp, 243bp, and 635bp as examples to illustrate the screening.

[0049] The sample processing method is the same as in Example 1.

[0050] 1. PCR amplification is as follows:

[0051] (1) Obtain the sample supernatant according to the processing method of feather marrow or blood sample described in Example 1, take 1.5 μ L as the template of PCR reaction, PCR reaction system (20 μ L): 1.5 μ l sample DNA template and 18.5 μ L PCR reaction reagent . Wherein the PCR reaction reagent comprises: primer pair, 2×PermixTaq (can adopt Tiangen Biotechnology Co., Ltd. reagent), double distilled water to form; Primer pair comprises upstream primer and downstream primer; In descr...

Embodiment 3

[0058] Example 3 Rapid sample lysate screening experiment

[0059] It is easy to promote in production by using pigeon feather non-destructive sampling method to carry out gender identification, but the effective nucleic acid part contained in feather is less, which brings great difficulty to effective nucleic acid amplification. The inventor explored a variety of feather-like lysates in the early stage, and finally found that the fast sample lysate of the present invention includes DNA extraction lysate A (NP40-PBS buffer solution, see Example 1 for configuration method) and lysate B (TE The buffer solution, see Example 1 for the configuration method, has the best lysis effect, can effectively meet the amplification test, and realize the feasibility of rapid molecular identification of gender. The rapid lysate of the present invention was compared with other lysates, using SEQ ID NO: 1 and SEQ ID NO: 2 as primers, respectively using hen pigeon feather samples to verify the te...

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Abstract

The invention provides a primer, a kit and a method for rapid identification of pigeon gender. The primer shown in the SEQ ID NO:1 and SEQ ID NO:2 has the advantages of high specificity, high accuracy, high resolution and the like in design of CHD gene sequence in pigeons; the amplification result is directly carried out by agarose gel electrohoresis, which provides higher identification so that the amplification result is intuitive and clear, and the pigeon gender can be easily distinguished. The invention further provides a molecular identification method of the pigeon gender by the aid of the primer. In the method, efficient nucleic acid lysate is further provided on the basis of the above the specific amplification of the primer, the process of obtaining DNA from feather pulp samples is greatly simplified, and time and economic cost in identification of the pigeon gender are reduced. The method and the kit for identification of pigeon gender with the PCR (polymerase chain reaction)-agarose electrophoresis technology and a genotyping method are simple in operation, rapid in identification, low in cost and abroad in the market utilization prospect.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a primer, a kit and a method for quickly identifying the sex of pigeons. Background technique [0002] Pigeons are monomorphic birds, and the appearance of male and female pigeons is very different. It is difficult to distinguish young pigeons from adult pigeons in terms of appearance or other behaviors. Sex identification is especially important in pigeon breeding and controlling breeding costs. If the sex ratio is inappropriate, it is not conducive to industrialized feeding management and breeding; individuals of different sexes have different nutritional needs, especially in the laying pigeon industry of the "double-female pairing" model. Therefore, early sex identification will help meet production needs and has important practical and economic value. [0003] At present, the traditional methods used for sex identification of pigeons mainly include anal inversion, l...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6888C12Q1/6879C12N15/11
Inventor 张莉
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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