TY - JOUR
T1 - A computer-driven approach to PCR-based differential screening, alternative to differential display
AU - Consalez, G. Giacomo
AU - Cabibbo, Andrea
AU - Corradi, Anna
AU - Alli, Cristina
AU - Sardella, Milena
AU - Sitia, Roberto
AU - Fesce, Riccardo
PY - 1999/2
Y1 - 1999/2
N2 - Motivation: Polymerase chain reaction (PCR)-based RNA fingerprinting is a powerful tool for the isolation of differentially expressed genes in studies of neoplasia, differentiation or development. Arbitrarily primed RNA fingerprinting is capable of targeting coding regions of genes, as opposed to differential display techniques, which target 3' non-coding cDNA. In order to be of general use and to permit a systematic survey of differential gene expression, RNA fingerprinting has to be standardized and a number of highly efficient and selective arbitrary primers must be identified. Results: We have applied a rational approach to generate a representative panel of high-efficiency oligonucleotides for RNA fingerprinting studies, which display marked affinity for coding portions of known genes and as shown 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 nucleotide databases, each containing the complete collection of deposited human or murine cDNAs. The simulation approach and experimental protocol proposed here permit the efficient isolation of coding cDNA fragments from differentially expressed genes. Availability: Freely available on request from the authors. Contact: fesce.riccardo@@@hsr.it.
AB - Motivation: Polymerase chain reaction (PCR)-based RNA fingerprinting is a powerful tool for the isolation of differentially expressed genes in studies of neoplasia, differentiation or development. Arbitrarily primed RNA fingerprinting is capable of targeting coding regions of genes, as opposed to differential display techniques, which target 3' non-coding cDNA. In order to be of general use and to permit a systematic survey of differential gene expression, RNA fingerprinting has to be standardized and a number of highly efficient and selective arbitrary primers must be identified. Results: We have applied a rational approach to generate a representative panel of high-efficiency oligonucleotides for RNA fingerprinting studies, which display marked affinity for coding portions of known genes and as shown 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 nucleotide databases, each containing the complete collection of deposited human or murine cDNAs. The simulation approach and experimental protocol proposed here permit the efficient isolation of coding cDNA fragments from differentially expressed genes. Availability: Freely available on request from the authors. Contact: fesce.riccardo@@@hsr.it.
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U2 - 10.1093/bioinformatics/15.2.93
DO - 10.1093/bioinformatics/15.2.93
M3 - Article
C2 - 10089194
AN - SCOPUS:0033007486
VL - 15
SP - 93
EP - 105
JO - Bioinformatics
JF - Bioinformatics
SN - 1367-4803
IS - 2
ER -