Corn leaves (V6 growth stage) from greenhouse grown plants
were collected and subsequently ground to a fine powder in liquid
nitrogen. RNA was extracted using E.Z.N.A. total RNA Kit II (Omega
Bio-tek Norcross, USA). The mRNA was enriched in the samples
through the use of Omega bio-tek's Mag-Bind mRNA enrichment
kit. The following changes were made to increase mRNA capture:
(1) The bead-to-sample ratio was increased from 0.5:1 to 1:1. (2)
The supernatant from the first mRNA capture step was rehybridized
in a second pass to increase the amount of mRNA captured. (3)
Washing steps were reduced from 3 times to 1. (4) The sample was
heated to 80 C for 2 min during the elution step. The mRNA was
subsequently concentrated using ethanol and sodium acetate to
precipitate the mRNA. The mRNA was resuspended in water,
quantified using a NanoDrop spectrophotometer, and stored
at -80 C.