T. H. (Henry) Hung [email protected], University of Oxford

📬 For any intended reader only

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<aside> ⚠️ HMW DNA is very prone to hydrodynamic shearing. Wide-bore tips are recommended in handling HMW DNA. Otherwise, pipette really slowly and handle very gently.

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Reagents

<aside> 💡 The protocol uses 1 g of fresh foliage. Scale up the amount of reagents when working with more samples.

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<aside> ☠️ Chloroform and β-mercaptoethanol are very toxic and volatile. Perform the experiment in the fumehood and dispose the waste properly.

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• [ ] 20 mL Carlson Lysis Buffer (Fisher Scientific 50-196-447) / CTAB Buffer (Promega MC1411)
  • Carlson Lysis Buffer Recipe
• [ ] 20 μL RNase A (Thermofisher 12091021)
• [ ] 20 mL Chloroform / Chloroform-isoamyl alcohol (24:1) (Sigma C0549-1PT) / Phenol-chloroform-isoamyl alcohol (25:24:1)
• [ ] 14 mL Isopropanol
• [ ] 4 mL 70% EtOH
• [ ] 50 μL β-mercaptoethanol
• [ ] 210 μL TE Buffer (Thermofisher 12090015)
• [ ] QIAGEN Blood and Cell Culture DNA Maxi Kit (Qiagen 13362)

Part 1: DNA extraction

⏱️ 1.5-hour hands-on time ⏱️ 1.5-hour procedure