A SIMPLE SUBTRACTION METHOD FOR THE ISOLATION OF CELL-SPECIFIC GENES USING MAGNETIC MONODISPERSE POLYMER PARTICLES

In this study we present a simple subtraction method for the isolation of cell-type-specific genes using magnetic beads. Biotinylated first- strand cDNA is generated from one cell type and immobilized onto magne tic streptavidin beads. Poly A(+) RNA, isolated from a different cell type by use of oligo-dT bends, is then hybridized to the immobilized c DNA. Beads with hybridized mRNA are subsequently removed from the solu tion by attraction to a magnet. The cell-specific mRNA, left in soluti on, is finally converted to a radiolabeled cDNA probe in order to scre en cDNA libraries. In this study we present an example of a successful subtraction strategy involving three cell types: the pre-B-cell line Reh; 12-O-tetradecanoylphorbol-13-acetate (TPA)-stimulated Burkitt's l ymphoma line Daudi; and TPA-stimulated T lymphocytes. This strategy wa s chosen due to our interest in gene products known to be expressed in TPA-stimulated B and T lymphocytes, but not in Reh cells. Several pre viously unknown genes were identified.