Generation of Full-Length cDNA Libraries Enriched for Differentially Expressed Genes

With sequence analysis of the human genome well underway, the scientific focus is shifting toward understanding the fundamentals of gene function. Sequence information alone is insufficient for a full understanding of gene function, expression, and regulation. Modern approaches such as subtractive hybridization (1 –5 ), differential display (6 ,7 ), serial analysis of gene expression (SAGE) (8 ), or complementary DNA (cDNA) arrays (9 ,10 ) allow researchers to monitor the differential expression of many genes in parallel. However, cDNAs identified by these techniques are often only short fragments. Further analyses, such as functional assays, require full-length cDNA clones. Two dominant approaches for cloning full-length cDNAs are cDNA library screening and PCR-based rapid amplification of cDNA ends (RACE) technology. Both techniques require knowledge of the cDNA sequence and are only useful for cloning a few genes at a time. Cloning full-length cDNAs for multiple genes and generation of full-length cDNA libraries enriched for differentially expressed genes in an efficient high-throughput manner still remain challenging.