Specification:
Principle: micro spin column (silica matrix)
Minimum elution volume: 10 μl
Sample size: ≤25 mg fixed tissue
Protocol: Isolation of DNA from paraffin-fixed tissue
Please Read Important Notes Before Starting Following Steps.
1. Add up to 25 mg of paraffin slice sample to a microcentrifuge tube.
2. Add 1 ml xylene and mix well. Close the lid and vortex vigorously for 10 secs. Incubate the sample at room temperature until
the paraffin is dissolved completely.
3. Centrifuge at full speed for 5 mins. Remove the supernatant by pipetting.
4. Add 1 ml ethanol (96~100%) to the deparaffined tissue and mix gently by vortexing.
5. Centrifuge at full speed for 3 mins. Remove the supernatant by pipetting.
6. Repeat step 4 and 5.
7. Incubate at 37°C for 10~15 mins to evaporate ethanol residue completely.
8. Add 200 μl FATG1 Buffer and mix well.
9. Add 20 μl Proteinase K to the sample mixture. Mix thoroughly by vortexing.
10. Incubate at 60°C until the tissue is lysed completely (1~3 hrs). Vortex occasionally during incubation.
-Sample can be incubated overnight as well for complete lysis.
11. (Optional) If RNA-free genomic DNA is required, add 4 μl of 100 mg/ml RNase A (not provided). Mix thoroughly by vortexing
and incubate at room temperature for 2 mins.
12. Incubate at 90°C for 30 mins. Vortex occasionally during incubation.
13. Add 200 μl FATG2 Buffer to the sample mixture, mix thoroughly by pulse-vortexing.
14. Add 200 μl ethanol (96~100%) to the sample mixture. Mix thoroughly by pulse-vortexing.
15. Place a TG Micro Column in a Collection Tube. Transfer the mixture carefully to the TG Micro Column. Centrifuge at full
speed (~18,000 xg) for 1 min then place the TG Micro Column to a new Collection Tube.
16. Add 400 μl W1 Buffer to the TG Micro Column. Centrifuge at full speed for 1 min then discard flow-through.
-Make sure that ethanol has been added into W1 Buffer at the first open.
17. Add 650 μl Wash Buffer to the TG Micro Column. Centrifuge at full speed for 1 min then discard flow-through.
-Make sure that ethanol has been added into Wash Buffer at the first open.
18. Centrifuge at full speed for an additional 3 mins to dry the column.
-Important Step! This step will remove the residual liquid.
19. Add ≥10 μl of preheated Elution Buffer or ddH2O (pH 7.5~9.0) to the membrane of the TG Micro Column. Stand the TG Micro
Column for 3 mins.
-Important Step! For effective elution, make sure that the elution solution is dispensed onto the membrane center and is
absorbed completely.
20. Centrifuge at full speed for 2 mins to elute DNA.
