Soil sample DNA extraction (SuperSoil)

Add 0.2mL ⌀=1.0 mm zirconia beads, 0.2mL ⌀=0.5 mm zirconia beads, 800µL SD1 solution, 0.5g Sample into a 1.5 ml microcentrifuge tube. Vortex briefly to mix.

Homogenize samples using Vortex-Genie® 2 with SI-H524 Horizontal microtube holder. Vortex at maximum speed for 0h 4m 0s

Centrifuge the 1.5 ml microcentrifuge tube at14000rpm,25°C

Add 200µL SD2 solution to a clean 1.5 ml microcentrifuge tube.

Transfer 500µL supernatant to the tube in the previous step. Vortex 0h 0m 5s for mixing.

Centrifuge at14000rpm,25°C

Add 600µL SD3 solution to a clean 1.5 ml microcentrifuge tube. Avoiding the pellet, transfer up to 700µL of supernatant from the previous step to that tube. Vortex 0h 0m 5s for mixing.

Take 650µL to spin column (with collection tube) and centrifuge at14000rpm,25°C

Discard the flow-through and repeat step 8 to ensure all the lysates have passed through the spin column.

Carefully place the spin column into a clean 2 ml collection tube. Avoid splashing any flow-through onto the spin column.

Add 500µL of Solution SEA to the spin column. Centrifuge at 14000rpm,25°C

Discard the flow-through and return the spin column to the same 2 ml collection tube.

Add 500µL 80% ethanol to the spin column. Centrifuge at 14000rpm,25°C

Discard the flow-through and place the spin column into a new 2 ml collection tube.

Centrifuge at 14000rpm,25°C Carefully place the spin column into the new 1.5 ml microcentrifuge tube

Add 50µL RNase-free water (DEPC-treated water) to the center of the white filter membrane.

Centrifuge at 14000rpm,25°C Discard the spin column. The DNA will be in the microcentrifuge tube.

Use BioTek Take3 & Take3 Trio Microvolume Plates to measure the quality and quantity of your specimens. And then upload them to Google Drive (1 MolecularWork (Direct edit) 分生實驗號碼 (直接編輯) w_ Primer list)