Strategy for chromosomal assignment of expressed sequences derived from heteronuclear RNA

Abstract

The chromosomal localization of transcribed sequences/genes is one of the objectives of the human genome project. Here, we describe a novel strategy for fast and dependable chromosomal assignment of expressed sequences that contain Alu sequences. Alu-PCR was performed on cDNA that was derived from heteronuclear (hn) RNA. hn-cDNA libraries are utilized for the identification of genes from extended human chromosomal regions or entire chromosomes. For chromosomal assignment, Alu-PCR products larger than 500 bp were hybridized against genomic DNA of somatic cell hybrids that was also amplified by Alu-PCR. Hybridization signals obtained within 2-3 h of exposure allow localization of cDNA-derived Alu-PCR products to single chromosomes. This technique is particularly useful for the analysis of cDNA libraries derived from hn-RNA. Using hn-cDNA clones from different chromosomes, we demonstrate the accuracy and reliability of the mapping strategy.