SuperSoil - Soil DNA Extraction
Yin-Tse Huang, Jie Hao Ou
Published: 2023-03-23 DOI: 10.17504/protocols.io.4r3l27864g1y/v1
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Abstract
SuperSoil - Soil DNA Extraction
Steps
Lysis
1.
Add following materials to 1.8 ml centrifuge tube
- 250 mg soil
- 1.5 g Bead
- 800 µl SD1
2. Vortex horizontally and ensure the cap is tightly sealed
Vortex for 0h 10m 0s at max speed
Note
3.
14000rpm
4. Expect 500–600 µl
Transfer the supernatant to a clean 1.5 ml or 2 ml tube
Note
Remove inhibitor
5. White precipitate forms.If too much AlCl3 (SD2) is added, it will cause all the DNA to be precipitated along with the inhibitory substances.See Mustafa (2017) for more info.Irfan Mustafa, Hadiatullah and Sustiyah 2017 Removal of humic acid from peat soils by using AlCl3 prior to DNA extraction AIP Conference Proceedings doi.org/10.1063/1.4983434
Add 200 ul SD2
Note
Citation
6.
Vortex for 0h 0m 5s at max speed
7. Don't worry if the supernatant is yellow.
14000rpm
Note
8. Expect 500–600 µl
Transfer the supernatant to a clean 1.5 ml or 2 ml tube
Note
Bind DNA to silica membrane
9.
Add 600 µl SD3
10.
Vortex for 0h 0m 5s at max speed
11.
Transfer 650 ul lysate to spin column
12.
14000rpm, discard flow-through
13.
Transfer remaining lysate to spin column
Wash silica membrane
14.
Add 500 ul SEA to spin column
15.
14000rpm, discard flow-through
16.
Add 500 ul S5 to spin column
17.
14000rpm, discard flow-through
18.
14000rpm, discard flow-through
Elution
19.
Add 50–100 ul S6
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