Capillary Gel Electrophoresis
Capillary Gel Electrophoresis (CGE) analysis of RNA integrity and purity
was performed with a 5200 Fragment Analyzer System (Agilent, CA, USA),
using the DNF-471 RNA Kit (15 nt) (Agilent, CA, USA). The capillary
cassette used was FA 12-Capillary Array Short, 33 cm (Agilent, CA, USA).
Samples were diluted to <100 ng/ul in nuclease free water.
Before each separation, a pre-run voltage was applied (8 kV for 30
seconds), the capillaries were conditioned with the conditioning
solution and the capillaries were dipped twice in the rinse buffer.
Following this, the capillaries were filled with RNA separation gel (by
pressure) and then the sample was introduced using a voltage injection
(5 kV for 4 seconds). The separation was then conducted by applying a
voltage of 8 kV for 45 minutes. Detection was carried out using Laser
Induced Fluorescence (LIF), by fluorescent dye tagging of the RNA.