The selective enrichment of rough endoplasmic reticulum fractions (RER) obtained from secretory tissues provides for a highly-efficient method for purifying mRNA molecules destined to be used as templates in the synthesis of secretory proteins. We have shown that the use of a non- ionic, low-viscosity linear gradient fractionates polysomes/free ribosomes and RER in distinct regions of the gradient, by Northern analysis in accordance with their predicted densities. Both mouse liver tissue and human liver cell lines have been tested and shown to segregate RER on density gradients. Differential mRNA display experiments were performed with RER bound mRNA cell lines versus healthy human liver tissue and total cell line mRNA as controls. PCR fragments differentially amplified in the cell line total and RER lanes, but not in the healthy patient control, were cloned and sequenced. No extensive homologies have been seen for most clones which we have sequenced so far. We are currently screening a human liver cDNA library to identify full- length clones.
