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Extraction method for environmental DNA of neophocaena phocaenoides asiaeorientalis and application of method

An extraction method, finless porpoise technology, applied in the field of molecular biology, can solve the problems of large weather influence factors, difficult long-term monitoring, labor and material resources consumption, etc., and achieve the effect of wide linear range, high efficiency and strong specificity

Pending Publication Date: 2018-10-09
FRESHWATER FISHERIES RES CENT OF CHINESE ACAD OF FISHERY SCI
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Problems solved by technology

The traditional species monitoring method uses visual and auditory methods to conduct on-the-spot surveys, which consumes a lot of manpower and material resources, and is greatly affected by the weather, making it difficult to achieve long-term monitoring. Therefore, it is urgent to adopt a new method to carry out the activities of the Yangtze finless porpoise Investigation of the scope and its regularity

Method used

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  • Extraction method for environmental DNA of neophocaena phocaenoides asiaeorientalis and application of method
  • Extraction method for environmental DNA of neophocaena phocaenoides asiaeorientalis and application of method
  • Extraction method for environmental DNA of neophocaena phocaenoides asiaeorientalis and application of method

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Experimental program
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Embodiment 1

[0025] 800mL of water samples were collected in a brand new 1L sealable jar, and 6 parallel samples were set up in the water area of ​​the Yangtze River South Pass (32°1′37.21″N, 120°26′50.68″E). In order to prevent large particles of sediment in the water sample from clogging the filter membrane, first filter the water sample with disposable medical gauze, and replace the gauze after filtration. Sampling equipment was disinfected with 10% bleach solution, rinsed with sterile water and dried before each use.

[0026] The samples were refrigerated and transported back to the laboratory, and 500 mL of water samples were filtered with a nylon filter membrane with a pore size of 10 μm. After suction filtration, the folded filter membrane was submerged in a 1.5mL EP tube filled with 95% ethanol and stored at -20°C.

[0027] DNA was extracted independently from the filter membrane of each water sample using E.Z.N.A.TM Water DNA Kit (OMEGA bio-tek, USA). After the filter membrane w...

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Abstract

The invention discloses an extraction method for environmental DNA of neophocaena phocaenoides asiaeorientalis and application of the method. The method comprises the following steps of collecting water samples of an area to be identified, carrying out filtration treatment, extracting the DNA by adopting kits, carrying out real time PCR amplification and determining whether the neophocaena phocaenoides asiaeorientalis exists in the area to be identified or not according to the peak value of a solubility curve. The method has the advantages of being high in efficiency, high in specificity, highin correlation coefficient, wide in linear range and the like; and whether the neophocaena phocaenoides asiaeorientalis exists in the surveyed water area can be rapidly and accurately identified.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and relates to a method for resource investigation or protection of germplasm resources, in particular to a method for extracting environmental DNA of Yangtze finless porpoise and its application. Background technique [0002] The Yangtze River finless porpoise (Neophocaena phocaenoides asiaeorientalis) is a small toothed whale listed as a national second-class protected animal. It is mainly distributed in the middle and lower reaches of the Yangtze River, Dongting Lake and Poyang Lake and their main tributaries. It is a relatively independent freshwater subspecies. According to the results of the international joint expedition of Yangtze River dolphins at the end of 2006, the number was about 1,800 at that time, and it continued to decline at a rate of at least 5% per year. Therefore, it is urgent to take further measures to protect wild life. To protect the Yangtze finless porpoise, i...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12Q1/6806C12Q1/686
CPCC12N15/1003C12Q1/6806C12Q1/686C12Q2561/113C12Q2563/107C12Q2545/114
Inventor 唐永凯吴昀晟俞菊华徐跑刘凯李建林李红霞于凡
Owner FRESHWATER FISHERIES RES CENT OF CHINESE ACAD OF FISHERY SCI
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