DNA EXTRACTION FROM AGAROSE |
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Last Update: December 2006 |
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Based on the kits developed by QIAGEN |
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| PROCEDURE | |
| 1. Excise DNA from agarose | |
| 2. Weigh gel slice and add 3 volumes of Buffer QG for every volume (100 mg = 100mL) of gel (to a maximum of 400 mg per column) | |
| 3. Incubate at 50oC for 10 min (until the gel slice has dissolved). Mix by vortexing to gelp dissolution (for >2% gels, increase incubation time) | |
4. The color of the mixture should still be yellow once the gel has dissolved (if not: add 10 mL of 3M sodium acetate, pH 5.0 and mix. The colors should turn yellow) |
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| 5. Add 1 gel volume of isopropanol to sample mix if gel fragment is <500 bp or >4 kb | |
| 6. Place a QIAquick column in a 2 mL tube | |
| 7. Apply sample to column and centrifuge for 1 min (up to 800 mL per column. If more, use more columns) | |
| 8. Discard FT and place column back in tube | |
9. Wash with 0.75 mL of Buffer PE and centrifuge for 1 min |
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| 10. Discard FT and centrifuge column for an additional 1 minute | |
| 12. Place column in a clean 1.5 mL tube, add 30-50 mL Buffer EB or dH2O, incubate at RT for 1 min, and centrifuge for 1 min | |
| DNA is ready for downstream applications | |