Extraction of DNA from Phyllosphere Microbiota
- First, 6.0 g of rubber leaves was transferred into a sterile conical
flask containing 100 mL 0.1 mol/L phosphate buffer solution (pH=7.0),
and then 100 μL Tween 80 was added.
- Each sample was shaken with ultrasonic oscillation for 8 minutes, and
then shaken at a speed of 200 rpm for 30 minutes at 25°C.
- After 2 minutes of ultrasonic oscillation, sterilized forceps were
used to remove the leaves on the super-clean worktable.
- The microorganisms in the conical flask were collected using suction
filtration with a 0.45-μm microporous membrane.
- DNA was extracted from each sample using the MP Biomedicals™ FastDNA®
Spin Kit for Soil (CA, USA).
- The concentration and purity of DNA were verified using the NanoDrop
2000C (Thermo Scientific, USA) to A260/A280 = 1.8–2.0, and stored at
−20°C until use.