title: Efficient targeted transcript discovery via array-based normalization of RACE libraries.
abstract: Rapid amplification of cDNA ends (RACE) is a widely used approach for transcript identification. Random clone selection from the RACE mixture, however, is an ineffective sampling strategy if the dynamic range of transcript abundances is large. To improve sampling efficiency of human transcripts, we hybridized the products of the RACE reaction onto tiling arrays and used the detected exons to delineate a series of reverse-transcriptase (RT)-PCRs, through which the original RACE transcript population was segregated into simpler transcript populations. We independently cloned the products and sequenced randomly selected clones. This approach, RACEarray, is superior to direct cloning and sequencing of RACE products because it specifically targets new transcripts and often results in overall normalization of transcript abundance. We show theoretically and experimentally that this strategy leads indeed to efficient sampling of new transcripts, and we investigated multiplexing the strategy by pooling RACE reactions from multiple interrogated loci before hybridization.
authors.affiliation: Grup de Recerca en Informàtica Biomèdica, Institut Municipal d'Investigació Mèdica/Universitat Pompeu Fabra, Dr. Aiguader 88, 08003 Barcelona, Spain.
authors.forename: Sarah
authors.initials: S
authors.lastname: Djebali
authors.forename: Philipp
authors.initials: P
authors.lastname: Kapranov
authors.forename: Sylvain
authors.initials: S
authors.lastname: Foissac
authors.forename: Julien
authors.initials: J
authors.lastname: Lagarde
authors.forename: Alexandre
authors.initials: A
authors.lastname: Reymond
authors.forename: Catherine
authors.initials: C
authors.lastname: Ucla
authors.forename: Carine
authors.initials: C
authors.lastname: Wyss
authors.forename: Jorg
authors.initials: J
authors.lastname: Drenkow
authors.forename: Erica
authors.initials: E
authors.lastname: Dumais
authors.forename: Ryan R
authors.initials: RR
authors.lastname: Murray
authors.forename: Chenwei
authors.initials: C
authors.lastname: Lin
authors.forename: David
authors.initials: D
authors.lastname: Szeto
authors.forename: France
authors.initials: F
authors.lastname: Denoeud
authors.forename: Miquel
authors.initials: M
authors.lastname: Calvo
authors.forename: Adam
authors.initials: A
authors.lastname: Frankish
authors.forename: Jennifer
authors.initials: J
authors.lastname: Harrow
authors.forename: Periklis
authors.initials: P
authors.lastname: Makrythanasis
authors.forename: Marc
authors.initials: M
authors.lastname: Vidal
authors.forename: Kourosh
authors.initials: K
authors.lastname: Salehi-Ashtiani
authors.forename: Stylianos E
authors.initials: SE
authors.lastname: Antonarakis
authors.forename: Thomas R
authors.initials: TR
authors.lastname: Gingeras
authors.forename: Roderic
authors.initials: R
authors.lastname: Guigó
mesh_terms: D017398:Alternative Splicing; D002891:Chromosomes, Human, Pair 21 / Q000235:genetics; D002892:Chromosomes, Human, Pair 22 / Q000235:genetics; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D005091:Exons; D020869:Gene Expression Profiling / Q000379:methods*; D015723:Gene Library; D015894:Genome, Human; D006801:Humans; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D020033:Protein Isoforms / Q000235:genetics; D012313:RNA / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic
chemical_list: D018076:DNA, Complementary; D020033:Protein Isoforms; D012313:RNA