two-component Enhancer-inhibitor transposon mutagenesis system for functional analysis of the Arabidopsis genome

A modified Enhancer-Inhibitor transposon system was used to generate a series of mutant lines by single-seed descent such that multiple I insertions occurred per plant. The distribution of original insertions in the population was assessed by isolating transposon-flanking DNA, and a database of inse...

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Bibliographic Details
Published in:The Plant cell 1999-10, Vol.11 (10), p.1853-1866
Main Authors: Speulman, E, Metz, P.L.J, Arkel, G. van, Lintel Hekkert, B. te, Stiekema, W.J, Pereira, A
Format: Article
Language:English
Subjects:
Alleles
Arabidopsis - genetics
Arabidopsis thaliana
Base Sequence
Biosynthesis
DNA
DNA Primers
DNA Transposable Elements
Enhancer Elements, Genetic - genetics
Gels
genetic transformation
Genetic transposition
Genome, Plant
Genomes
Genomics
Genomics Articles
inhibitor tagged sites
insertion site sequences
insertional mutagenesis
insertions
Mutagenesis, Insertional
mutants
mutation
nucleotide sequences
Phenotypes
Plants
Polymerase Chain Reaction
Repressor Proteins - genetics
screening
transgenic plants
Transposons
Zea mays
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Summary:A modified Enhancer-Inhibitor transposon system was used to generate a series of mutant lines by single-seed descent such that multiple I insertions occurred per plant. The distribution of original insertions in the population was assessed by isolating transposon-flanking DNA, and a database of insertion sites was created. Approximately three-quarters of the identified insertion sites show similarity to sequences stored in public databases, which demonstrates the power of this regimen of insertional mutagenesis. To isolate insertions in specific genes, we developed three-dimensional pooling and polymerase chain reaction strategies that we then validated by identifying mutants for the regulator genes APETALA1 and SHOOT MERISTEMLESS. The system then was used to identify inserts in a class of uncharacterized genes involved in lipid biosynthesis; one such insertion conferred a fiddlehead mutant phenotype.
ISSN:1040-4651
1532-298X
DOI:10.1105/tpc.11.10.1853