ENRICHMENT OF CHROMOSOME-SPECIFIC HNCDNAS BY MAGNETIC BEAD COUPLED ALU SEQUENCES

We have employed a strategy for the rapid enrichment of cDNA clones fr om human chromosome 22 utilizing magnetic beads. Starting from a somat ic cell hybrid which retains chromosome 22 in rodent background, heter onuclear (hn) RNA was transcribed into hncDNA using poly dT-primers. U sing linker specific primers hncDNA was amplified by PCR. To identify chromosome 22 specific hncDNAs a highly human specific Alu consensus s equence (PD39) was biotinylated and hybridized to the PCR product of t he hncDNAs in solution. Hybridized hncDNA-PD39 complexes are captured using streptavidin-coated magnetic beads. Hybridized hncDNAs are selec tively amplified by PCR. To verify the chromosome specificity the hncD NA was used as probe for in situ hybridization. Following two rounds o f selection with magnetic beads there was an increasingly strong hybri dization signal on chromosome 22. The capturing of hncDNAs by magnetic beads as described in this study is faster and more efficient than pr eviously described methods for the isolation of chromosome specific hn cDNAs. The novel approach has been employed to generate hncDNAs highly enriched for chromosome 22 specific sequences.