Sacalia bealei microsatellite marked specific primer and detection method

A technology of microsatellite markers and terrapins, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., can solve problems such as unseen terrapins and achieve rich polymorphism Effect

Inactive Publication Date: 2014-03-12
INST OF ZOOLOGY GUANGDONG ACAD OF SCI
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  • Sacalia bealei microsatellite marked specific primer and detection method
  • Sacalia bealei microsatellite marked specific primer and detection method
  • Sacalia bealei microsatellite marked specific primer and detection method

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Embodiment 1

[0031] 1. Extraction of Genomic DNA from Spotted Terrapins

[0032] 1. Reagent preparation:

[0033] Digestion buffer: Weigh Tris0.6057g, EDTA18.612g, SDS2.5g, dilute to 500mL with sterilized ultrapure water (final concentration is Tris10mmol / L, EDTA0.1mol / L, SDS0.5%), adjust pH value to 8.0, store at room temperature for later use.

[0034] 2. DNA extraction:

[0035] Cut the tail muscle tissue of 33 spotted water turtles into 2mL centrifuge tubes, cut the tissue into pieces, add 800μL digestion buffer, 10μL proteinase K (20mg / mL), mix well and digest at 55℃ for 8-10h; After clearing, add 800 μL of saturated phenol, mix gently by inversion for 20 minutes, centrifuge at 10,000 g for 10 minutes, and extract the supernatant; then add 400 μL of saturated phenol, 400 μL of chloroform / isoamyl alcohol (24:1), and mix by inversion for 20 minutes. Centrifuge at 10000g for 10min, extract the supernatant; then add 800μL chloroform / isoamyl alcohol (24:1), mix gently by inversion for 2...

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Abstract

The invention discloses a Sacalia bealei microsatellite marked specific primer and a detection method. The detection method comprises the following steps: performing polymerase chain reaction (PCR) amplification on microsatellite site sequence by taking Sacalia bealei genome DNA as a template and using the microsatellite marked specific primer; performing non-denaturing polyacrylamide gel electrophoresis detection on the PCR amplification products; and performing polymorphic analysis on microsatellite sites according to the detection result. The Sacalia bealei microsatellite marked specific primer and the detection method thereof, provided by the invention, can be applied in the fields of genetic diversity level evaluation, population genetic structure analysis and relationship identification of Sacalia bealei.

Description

technical field [0001] The invention belongs to the technical field of molecular markers, and in particular relates to a specific primer and a detection method for microsatellite markers of spotted turtles. Background technique [0002] The eye-spotted water turtle (Sacalia bealei) belongs to the genus Eye-spotted turtle of the order Turtles and is endemic to China. It is mainly distributed in Guangdong, Guangxi, Fujian, Anhui, Jiangxi, Guizhou, Hong Kong and other places, and lives in mountain streams. The spotted water turtle has edible, medicinal and ornamental value, and is an important economic freshwater turtle. In recent years, due to overhunting, trade, and habitat destruction, the resources of wild spotted water turtles have been decreasing. In order to scientifically protect the germplasm resources of spotted water turtles and realize the sustainable use of resources, the genetic diversity analysis of this species has been carried out. And artificial breeding is o...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6888C12Q2600/156
Inventor 龚世平李伟业滑留帅
Owner INST OF ZOOLOGY GUANGDONG ACAD OF SCI
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