A method for the elimination of false positives generated by the mRNA differential display technique.
The recently described mRNA differential display method provides an attractive tool for the isolation of genes showing regulated expression in a variety of systems. A key step in this technique consists of the isolation of PCR-synthesized radioactive cDNAs corresponding to differentially expressed mRNAs. Here, we show that the purified cDNAs remain contaminated with unrelated cDNA sequences that may lead to the artifactual isolation of false positives in the subsequent steps of the method. A powerful assay for the detection and elimination of this contaminating material, allowing the specific isolation of clones corresponding to the regulated genes identified by the differential display, is provided.
Callard
D
D
Laboratoire de Biologie Moléculaire des Relations Plantes-Microorganismes CNRS-INRA (UMR CNRS 05), Castanet-Tolosan, France.
Lescure
B
B
Mazzolini
L
L
D017360:Arabidopsis / Q000235:genetics; D001345:Autoradiography; D015152:Blotting, Northern; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000302:isolation & purification*; D004340:Drug Contamination; D005189:False Positive Reactions; D015870:Gene Expression; D010957:Plasmids; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000032:analysis*; D012194:RNA-Directed DNA Polymerase
D018076:DNA, Complementary; D012333:RNA, Messenger; D012194:RNA-Directed DNA Polymerase