According
to the manufacturer’s instructions, 1 ml Trizol solution
was added to each frozen sample, and the lysate was aspirated
through a 20-gauge needle before centrifugation at 10,000g
for 3 min at room temperature.
Thereafter, all lysates were
diluted 1:1 with 70% ethanol and subjected to a mini-column
separation. After the binding of the RNA to the column,
DNA digestion was performed by using RNAse-free DNAse
(340 Kunitz units/ml) for 15 min at room temperature. After
the column had been washed three times, the RNA was
eluted with 30 μl RNAse-free water.