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Chemical regulated production of cDNAs from genomic DNA fragments in plants
Author(s) -
Møller Simon Geir,
Chua NamHai
Publication year - 2002
Publication title -
the plant journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.058
H-Index - 269
eISSN - 1365-313X
pISSN - 0960-7412
DOI - 10.1046/j.1365-313x.2002.01436.x
Subject(s) - complementary dna , genomic dna , biology , transgene , genetics , computational biology , exon , cdna library , intron , genomic library , arabidopsis , dna , gene , microbiology and biotechnology , base sequence , mutant
Summary We have developed a new chemical inducible genetic system that allows for the isolation of any cDNA molecule from in vitro generated genomic transgenes in transgenic plants. This system, termed regulated in vivo cDNA generation (RIDE), permits both targeted isolation of individual full‐length cDNA molecules and random isolation of any partial or full‐length cDNA from in planta genomic libraries. The RIDE system makes use of the 17‐β estradiol‐inducible promoter system linked to intron donor and acceptor sites in a new binary vector configuration. In transgenic Arabidopsis and tobacco plants, we show that the RIDE system can isolate low‐abundance full‐length cDNAs previously unattainable by conventional means at high efficiencies (75–85%). The ability to randomly isolate individual exons and exons spliced together from genomic libraries in planta suggest that this system can be used for the isolation of any cDNA molecules. The RIDE system thus appears to be an efficient and versatile system for the generation of potentially any cDNA molecule. Moreover, the ORF structural data generated will be of value in both verifying and correcting computational ORF predications in the databases available to the scientific community.