This reagent is a DNA extraction phenol reagent saturated with Tris-HCl (pH 8.0). It appears as a light yellow, clear liquid with a pungent odor. The upper layer consists of Tris-HCl buffer with pH > 7.8, while the lower layer is the phenolic phase, which contains 8-hydroxyquinoline as an antioxidant. This product is suitable for DNA isolation.
This reagent is for research use only and is not intended for clinical diagnosis or other purposes.
Materials to Prepare
Tissue Lysis Buffer
Tris-HCl (1M, pH 8.0)
Chloroform
Isoamyl alcohol
Isopropanol
75% Ethanol
TE Buffer
Procedure (For Reference Only)
Take an appropriate amount of Tissue Lysis Buffer and follow the steps below for DNA extraction:
1. Adjust pH: Ensure the pH of the lysis buffer is around 8.0 (add 0.1 volume of 1 mol/L Tris-HCl, pH 8.0 for adjustment if necessary).
2. Organic Extraction: Sequentially add the following reagents:
- 0.5 volume of DNA Phenol Reagent (Note: Allow the reagent to stand before use; use the lower organic phase. The upper layer is the Tris-HCl solution, which prevents oxidation of the lower organic phase).
- 0.5 volume of Chloroform:Isoamyl alcohol (24:1)
Vortex for 10 seconds to mix thoroughly, then incubate on ice for 15 minutes.
3. Centrifugation and Separation: After centrifugation, carefully transfer the upper aqueous phase to a new centrifuge tube.
4. DNA Precipitation: Add an equal volume of isopropanol, mix well, and precipitate the DNA.
5. Washing and Dissolution: Wash the precipitate once with 75% ethanol, air-dry briefly, and dissolve in an appropriate volume of TE buffer for subsequent use.
Precautions
1. DNA Phenol Reagent is highly corrosive. Avoid skin contact or inhalation. If the reagent turns red or brown, it indicates oxidation and should not be used.
2. For your safety and health, please wear a lab coat and disposable gloves during operation.
3. Use the reagent as soon as possible after opening to prevent affecting subsequent experimental results.
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