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Multi-polymerase chain reaction (PCR) primer and method for identifying clouded leopard and application of primer or method in identification of clouded leopard

A multiple, clouded leopard technology, applied in biochemical equipment and methods, recombinant DNA technology, microbial measurement/inspection, etc., can solve the problems of time-consuming, labor-intensive and costly, reduce the difficulty of sampling, economical and practical identification, simple and easy The effect of identification

Inactive Publication Date: 2014-09-24
ANHUI NORMAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] In view of the above-mentioned prior art, the purpose of the present invention is to overcome the limitation of measuring the clouded leopard species by morphological characteristics in the prior art, or the time-consuming and labor-intensive method of determining the species by measuring the DNA in feces by conventional methods And cost a lot of problems, provide a simple, quick and economical multiplex PCR primers for identifying clouded leopards and a method for identifying clouded leopards, and their application in identifying clouded leopards

Method used

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  • Multi-polymerase chain reaction (PCR) primer and method for identifying clouded leopard and application of primer or method in identification of clouded leopard
  • Multi-polymerase chain reaction (PCR) primer and method for identifying clouded leopard and application of primer or method in identification of clouded leopard
  • Multi-polymerase chain reaction (PCR) primer and method for identifying clouded leopard and application of primer or method in identification of clouded leopard

Examples

Experimental program
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Effect test

Embodiment 1

[0035] Put 0.5 g of the sample to be tested numbered 3 in Table 1 in a centrifuge tube and cut the sample to be tested in the centrifuge tube with sterilized scissors, then add 500 μL of lysate and 30 μL of 10% ten Sodium dialkyl sulfate and 3 μL of 20 mg / ml proteinase K were thoroughly mixed and then digested in a water bath at 56°C for 12 hours until the liquid in the tube was clear. Add 500 μL of Tris-balanced phenol to the above-mentioned digested mixture, shake slightly for 5 minutes, then place it in a centrifuge at 11000 r / min for 10 minutes, and take the supernatant after centrifugation. Repeat the above centrifugation step twice. Add 1,000 μL of frozen absolute ethanol to the supernatant obtained after repeated centrifugation, place it at -20°C for 1 hour, then centrifuge it in a centrifuge at 12,000 r / min for 13 minutes, discard the supernatant, and add 800 μL of 70% ethanol was shaken slightly for 0.5 min, then placed in a centrifuge at 13000 r / min for 13 min, and ...

Embodiment 2

[0038] Operate according to the method of Example 1, the difference is that the sample number to be tested is 4 (in Table 1), the consumption of each primer is 0.75 μ L, the consumption of the total DNA is 45 ng, and the annealing temperature is 55°C, the annealing time is 40s, the electrophoresis results are as follows figure 1 Shown (lane number is 4).

Embodiment 3

[0040] Operate according to the method of Example 1, the difference is that the sample number to be tested is 5 (in Table 1), the consumption of each primer is 1.25 μ L, the consumption of the total DNA is 55 ng, and the annealing temperature is 60°C, the annealing time is 25s, the electrophoresis results are as follows figure 1 Shown (lane number is 5).

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Abstract

The invention discloses a multi-polymerase chain reaction (PCR) primer for identifying a clouded leopard. The multi-PCR primer comprises primer pairs 1 shown in SEQ ID No:1 and SEQ ID No:2, and primer pairs 2 shown in SEQ ID No:3 and SEQ ID No:4. The invention also discloses a method for identifying the clouded leopard by using multi-PCR. The method comprises the following steps: extracting total deoxyribonucleic acid (DNA) of a to-be-tested sample and carrying out PCR amplification on the total DNA; and carrying out agarose gel electrophoresis detection on the amplified product, wherein the multi-PCR primer is the multi-PCR primer, and judging the to-be-tested sample to be the sample from the clouded leopard if two stripes appear in a lane. The invention also discloses an application of the multi-PCR primer or identification method in identification of the clouded leopard. The target of simply, feasibly, economically and practically identifying the clouded leopard is achieved by designing two pairs of primers.

Description

technical field [0001] The invention relates to the field of biological detection, in particular to a multiple PCR primer for identifying clouded leopards, a method for identifying clouded leopards using the multiple PCR primers, and their application in identifying clouded leopards. Background technique [0002] Accurate species identification based on taxonomy is a necessary prerequisite for human cognition of nature and sustainable development. It provides a knowledge and theoretical basis for the conservation of biodiversity, the sustainable use of species resources, and the development of new products from biological sources. For the identification of species, traditional methods mainly use morphological evidence. However, some shortcomings of this method have also caused limitations in the protection of endangered animals, such as phenotypic plasticity and genetic variability, inability to distinguish cryptic taxa, and restrictions by biological sex and developmental ...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/686C12Q2537/143C12Q2565/125
Inventor 晏鹏晏龙耿章珍吴孝兵
Owner ANHUI NORMAL UNIV
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