INS-1-EGFP or INS-1-EGFP-L10a stable cell lines were induced with doxycycline (Dox), treated with cycloheximide, washed with PBS, and lysed with 20mM HEPES [pH 7.4], 150mM KCL, 2M MgCl2, 1% NP-40. Lysates were homogenized in ice-cold polysome extraction buffer and homogenates clarified. Immunoaffinity-purification of polysomal RNA used goat anti-GFP (Nathaniel Heintz, Rockefeller Univ.), which was precipitated, resuspended, and quantified for DNA microarray experiments (See Supp methods for further details).
