GENOMIC DNA PREPARATION FROM PLANT TISSUES |
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Last Update: December 2006 |
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| PREPARE SOLUTIONS | |
| 2X CTAB buffer (10 mL): | Mix 2.8 mL of 5M NaCl, 0.2 g of CTAB (hexadecyltrimethylammonium bromide), 100 mL of b-mercatoethanol, 1 mL of 1M Tris-HCl pH 8, dH2O to 10 mL. Incubate at 65oC for the CTAB to go into solution |
| PROCEDURE | |
| 1. Harvest plant material and freeze in liquid nitrogen | |
| 2. Grind tissue (optional: freeze-dry tissues to aid in grinding - the more the tissue is made into a powder, the better the yield) to a fine powder | |
| 3. Add ~5 mL of fresh 2X CTAB buffer per ~1 g of tissue. Incubate at 65oC for 30 mins | |
4. Extract 2X with an equal volume of chlorophorm:isoamyl alcohol (24:1) |
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| 5. Precipitate with 0.8 volume of isopropanol / 0.1 volume of 3M NaoAc. Optional: leave ON at -20oC | |
| 6. Pellet at 10,000 rpms for 10-30 mins and wash pellet with 70% ethanol at RT | |
| 7. Resuspend pellet in TE buffer (with 10-20 mg/mL RNAse) | |