A computer-driven approach to PCR-based differential screening, alternative to differential display

Motivation: Polymerase chain reaction (PCR)-based RNA fingerprinting is a p owerful tool for the isolation of differentially expressed genes in studies of neoplasia, differentiation or development. Arbitrarily primed RNA finge rprinting is capable of targeting coding regions of genes, as opposed to di fferential display techniques, which target 3' non-coding cDNA. In order to be of general use and to permit ct systematic survey of differential gene expression, RNA fingerprinting has to be standardized and a number of highl y efficient and selective arbitrary primers must be identified. Results: We have applied a rational approach to generate a representative p anel of high-efficiency oligonucleotides for RNA fingerprinting studies, wh ich display marked affinity for coding portions of known genes and, as show n by preliminary results, of novel ones. The choice of oligonucleotides was driven by computer simulations of RNA fingerprinting reverse transcriptase (RT)-PCR experiments, performed on two custom-generated non-redundant nucl eotide databases, each containing the complete collection of deposited huma n or murine cDNAs. The simulation approach and experimental protocol propos ed here permit the efficient isolation of coding cDNA fragments from differ entially expressed genes.