The Preparation of Biologically Active Messenger RNA from Human Postmortem Brain Tissue

Abstract

MRNA was extracted from human postmortem brain tissue by alkaline phenol extraction of polysomes followed by oligo (dT)-cellulose chromatography. The mRNA preparations stimulated protein synthesis in a cell-free system containing wheat germ homogenate. The products of protein synthesis were analyzed by 1- and 2-dimensional gel electrophoresis. Numerous polypeptides, including tubulin subunit and actin isomers, were synthesized by the human mRNA. The MW range of polypeptides synthesized by human mRNA fractions from 2 brain specimens were identical, and analysis by 2-dimensional gel electrophoresis indicated qualitatively similar products. The yield of mRNA extracted per gram of human tissue was less than the yield obtained with rat forebrains from animals sacrificed immediately before brain removal and mRNA purification. A decrease in the amount of polysomes isolated from human tissue relative to rat brain tissue was a major factor contributing to the low yield. The MW distribution of polypeptides synthesized by human and rat brain mRNA fractions in wheat germ homogenate was similar; there was no indication for selective breakdown or inactivation of high MW mRNA species in the human tissue. It is possible to utilize postmortem tissue for molecular biological investigations of human brain mRNA.