Method for amplifying o complete sequence of mitochondria of golden coin turtle by using DNA extracted from nails
A technique of three-line box turtle and mitochondria
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[0029] The method provided in this example to amplify the full-length mitochondria of the box turtle using toenails to extract DNA comprises the following steps:
[0030] (1) Toenail sampling
[0031] Use surgical scissors to cut the toenails of the box turtle, soak them in absolute ethanol and store them at -20°C until use;
[0032] (2) Extraction of genomic DNA
[0033] Before DNA extraction, toenails were soaked in 1×PBS buffer at 4°C overnight, and washed 2-3 times during this period. Take about 10-20 mg of each toenail sample, and use the MicroElute Genomic DNA Kit (OMEGA, USA) kit to extract DNA. ① Add the toenail sample to a 1.5mL centrifuge tube, and add 210-220 μL of TLBuffer; ② Cut the toenail sample as much as possible , add 20 μL of OB solution; ③ place in a shaker air bath with a temperature of 60-70 rpm / min, digest at 56-60 ° C for 3-5 hours; ④ centrifuge at 20,000xg for 2 min, and take the supernatant; Buffer, mix in 70°C water bath for 10 minutes; ⑥Add 220 μ...
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