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Genomic DNA Isolation Protocol for 40 l Avian Whole Blood


AquaPure Genomic DNA Isolation Kit
Catalog #732-6340
Expected yield range: 100300 g DNA


Method
Cell Lysis
1. Use fresh blood within 10 sec of venipuncture to avoid clotting. Alternatively, collect whole blood in EDTA to prevent the blood from clotting and reduce DNA degradation. Blood may be either fresh or frozen. Store fresh blood sample at 28C for not more than 5 days to obtain optimal DNA isolation results.

2. Add 40 l whole blood to a 15 ml microfuge tube containing 6 ml cell lysis solution. Quickly pipet up and down 35 times to lyse the cells. Usually no incubation is required; however, if cell clumps are visible after mixing, incubate at 37C until the solution is homogeneous. Samples are stable in cell lysis solution for at least 18 months at room temperature.

RNase Treatment (Optional)
1. Add 30 l RNase A solution to the cell lysate.

2. Mix the sample by capping the tube and inverting it 25 times, then incubate at 37C for 1560 min.

Protein Precipitation
1. Cool sample to room temperature.

2. Add 2 ml protein precipitation polution to the RNase A treated cell lysate.

3. Vortex vigorously at high speed for 20 sec to mix the protein precipitation solution uniformly with the cell lysate.

4. Centrifuge at 2,000 x g for 10 min. The precipitated proteins will form a tight pellet. If the protein pellet is not visible or is not tight, repeat step 3, incubate on ice for 5 min, then repeat step 4.

DNA Precipitation
1. Leaving behind the precipitated protein pellet, pour the supernatant containing the DNA into a 15 ml centrifuge
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