PCR method for rapid identification of four types of blue crabs of scylla
A mud crab, rapid technology, applied in the fields of biochemical equipment and methods, microbial determination/inspection, etc., can solve the problems of poor labeling repeatability, difficult to master, low identification accuracy, etc., to achieve simple operation, improve efficiency and The effect of accuracy
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[0041] 1. Extraction of Genomic DNA of 4 Scyllas
[0042]Cut a small amount of muscle tissue (about 100-150 mg) from blue crabs and place it in 500 μl tissue extract solution (10 mmol / L Tris-CI, pH 8.0; 100 mmol / L LEDTA, pH 8.0; 100 mmol / L NaCl; 0.5% SDS) Cut into pieces, add proteinase K (final concentration 20mg / ml) and RNase A (final concentration 100μg / ml), mix thoroughly, digest at 55°C for 2-3 hours until clear; mix with phenol: chloroform: isoamyl alcohol solution (volume ratio 25:24:1) twice; then precipitate DNA with 2 volumes of absolute ethanol, wash with 70% ethanol and dry naturally, then dissolve in 50 μL TE (10 mmol / L Tris-HCl, pH8.0; 10mmol / L EDTA, pH8.0); Finally, the DNA concentration was diluted to 100ng / μl and stored at -20°C for later use;
[0043] 2. Mitochondrial COI gene sequence analysis and species-specific primer design
[0044] Download the COI sequences of four kinds of blue crabs from the public database GenBank, use DNAMAN 4.0 biological softwa...
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