RNA Isolation from Diverse Biological Samples

RNA was isolated from maternal and fetal plasma, CSF, and amniotic fluid using the Viral Total Nucleic Acid Purification Kit (Promega, Madison, WI) on a Maxwell 48 RSC instrument (Promega, Madison, WI) as previously reported [23 (link)]. Fetal and maternal tissues were processed with RNAlater (Invitrogen, Carlsbad, CA) according to the manufacturer’s protocols. RNA was recovered from tissue samples using a modification of a previously described method [24 (link)]. Briefly, up to 200 mg of tissue was disrupted in TRIzol (Life Technologies, Carlsbad, CA) with 2 x 5 mm stainless steel beads using a TissueLyser (Qiagen, Germantown, MD) for 3 minutes at 25 r/s for 2 cycles. Following homogenization, samples in TRIzol were separated using Bromo-chloro-propanol (Sigma-Aldrich, St. Louis, MO). The aqueous phase was collected, and glycogen was added as a carrier. The samples were washed in isopropanol and ethanol precipitated. RNA was re-suspended in 5 mM Tris pH 8.0 and stored at -80°C.