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

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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 spec

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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Example Embodiment

[0034] Example 1

[0035] Put 0.5g of the sample numbered 3 in Table 1 in a centrifuge tube and cut the sample in the centrifuge tube with sterile scissors, and then add 500μL of lysate to the centrifuge tube, 30μL of 10% ten Sodium dialkyl sulfate and 3μL of 20mg / ml proteinase K were thoroughly mixed and digested in a water bath at 56°C for 12h until the liquid in the tube was clear. Add 500 μL of Tris-balanced phenol to the digested mixture, shake it slightly for 5 minutes, and centrifuge for 10 minutes on a 11000r / min centrifuge, and take the centrifuged supernatant. Repeat the above centrifugation step twice. Add 1000 μL of frozen absolute ethanol to the final supernatant after repeated centrifugation and place it at -20°C for 1 hour, then place it on a centrifuge at 12000r / min for 13 minutes, discard the supernatant, and add to the resulting pellet 800 μL of 70% ethanol, and gently shake for 0.5 min, place it in a centrifuge at 13000 r / min and centrifuge for 13 min, and di...

Example Embodiment

[0037] Example 2

[0038] Operate according to the method of Example 1, except that the sample number to be tested is 4 (in Table 1), the amount of each primer is 0.75 μL, the amount of total DNA is 45 ng, and the annealing temperature is 55℃, the annealing time is 40s, the electrophoresis result is as follows figure 1 As shown (lane number 4).

Example Embodiment

[0039] Example 3

[0040] The operation was carried out according to the method of Example 1, except that the sample number to be tested was 5 (in Table 1), the amount of each primer was 1.25 μL, the amount of total DNA was 55 ng, and the annealing temperature was 60℃, the annealing time is 25s, the electrophoresis result is as follows figure 1 As 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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