The study of expressed genes has had a great impact on biological research (1). Expressed genes are the basic functional units of genomic DNA. Because these regions cannot be identified from genomic sequence information per se, the gene's products, messenger RNAs or proteins, must be isolated from cells, directly sequenced, and identified. As we steadily build up sizable expression databases that currently include more than 92,000 of the roughly 100,000 total human genes, the process of identifying the remaining undiscovered genes is becoming progressively more difficult. Existing databases of expressed genes now include virtually all of the abundantly expressed human genes—the easier to reach “low hanging fruit on the tree,” as well as many middle and rarely expressed genes. The genes that are still undiscovered are expressed at low levels or are specifically expressed only in certain cell types, developmental stages, or growth conditions. Such genes hold the promise of including key regulatory factors responsible for differentiated phenotypes, developmental progression, or cell growth regulation. As we move forward to identify these genes, highly efficient methods of removing, i.e., subtracting, the bulk of identified, abundant genes from cDNA libraries are required.