IDENTIFICATION OF CHROMOSOME-SPECIFIC SEQUENCE-TAGGED SITES BY ALU-PCR

Recently, methods have been developed for the isolation of expressed s equences from particular human chromosomes. Using Alu consensus sequen ces as primers, cDNA synthesis has been initiated from interspecies hy brid cell lines that contain single human chromosomes. Alu consensus s equences have also been utilized to amplify human genomic sequences vi a polymerase chain reaction (PCR). Here, we describe the use of Alu-PC R to isolate expressed sequences from human chromosomes selectively. H eteronuclear (hn) RNA is transcribed into cDNA by using poly-(dT)15 pr imer sequences. Subsequently, human specific cDNA sequences are amplif ied by Alu-PCR and cloned into pBluescript. To verify the chromosomal assignment, cloned PCR products are sequenced, converted into STS mark ers, and tested on a different somatic hybrid that contains human chro mosome 22. The method provides a fast, reliable way to identify expres sed sequence tagged sites from selected human chromosomes.