title: Quantitative cDNA-AFLP analysis for genome-wide expression studies.
abstract: An improved cDNA-AFLP method for genome-wide expression analysis has been developed. We demonstrate that this method is an efficient tool for quantitative transcript profiling and a valid alternative to microarrays. Unique transcript tags, generated from reverse-transcribed messenger RNA by restriction enzymes, were screened through a series of selective PCR amplifications. Based on in silico analysis, an enzyme combination was chosen that ensures that at least 60% of all the mRNAs were represented by an informative sequence tag. The sensitivity and specificity of the method allows one to detect poorly expressed genes and distinguish between homologous sequences. Accurate gene expression profiles were determined by quantitative analysis of band intensities, and subtle differences in transcriptional activity were revealed. A detailed screen for cell cycle-modulated genes in tobacco demonstrates the usefulness of the technology for genome-wide expression analysis.
authors.affiliation: Department of Plant Systems Biology, Flanders Interuniversity Institute for Biotechnology, Ghent University, K.L. Ledeganckstraat 35, Belgium.
authors.forename: P
authors.initials: P
authors.lastname: Breyne
authors.forename: R
authors.initials: R
authors.lastname: Dreesen
authors.forename: B
authors.initials: B
authors.lastname: Cannoot
authors.forename: D
authors.initials: D
authors.lastname: Rombaut
authors.forename: K
authors.initials: K
authors.lastname: Vandepoele
authors.forename: S
authors.initials: S
authors.lastname: Rombauts
authors.forename: R
authors.initials: R
authors.lastname: Vanderhaeghen
authors.forename: D
authors.initials: D
authors.lastname: Inzé
authors.forename: M
authors.initials: M
authors.lastname: Zabeau
mesh_terms: D001483:Base Sequence; D017931:DNA Primers / Q000494:pharmacology; D018076:DNA, Complementary / Q000378:metabolism; D005821:Genetic Techniques; D016678:Genome; D008957:Models, Genetic; D008969:Molecular Sequence Data; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction; D011110:Polymorphism, Genetic; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics
chemical_list: D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D012313:RNA