Large scale cDNA sequencing of 3'-directed cDNA libraries allows the collection of hundreds of novel sequences (gene signatures) and understanding of their expression levels in various tissues. Granulocytoid cells were induced from the human promyelocytic leukemia cell line 60 (HL60) with dimethyl sulfoxide (DMSO), and more than 1000 gene signatures were collected from a 3'-directed cDNA library of the granulocytoid cells. By selecting appropriate sequences for primer design, we systematically assigned chromosomes of these novel gene signatures by means of the polymerase chain reaction. We used a monochromosomal hybrid cell panel DNA as the template and localized 155 gene signatures to individual chromosomes. The results showed that 125 of them were assigned to single chromosomes and 30 to more than one. Genes assigned to more than one chromosome tend to be expressed ubiquitously, and their sequences are often conserved among humans and rodents. A highly sensitive laser fluorescent analyzer and ethidium staining were used for simple and large-scale PCR mapping.