Ac/Ds transposon mutagenesis in Arabidopsis thaliana: mutant spectrum and frequency of Ds insertion mutants

Abstract

Using a two-component Ac/Ds system consisting of a stabilized Ac element (Acc1) and a non-autonomous element (DsA), 650 families of plants carrying independent germinal DsA excisions/transpositions were isolated. Progenies of 559 of these Acc1/DsA families, together with 43 families of plants selected for excision/transposition of wild-type (wt) Ac, were subjected to a broad screening program for mutants exhibiting visible alterations. This resulted in the identification of 48 mutants showing a wide variety of mutant phenotypes, including embryo lethality (24 mutants), chlorophyll defects (5 mutants), defective seedlings (2 mutants), reduced fertility (5 mutants), reduced size (3 mutants), altered leaf morphology (2 mutants), dark green, unexpanded rosette leaves (3 mutants), and aberrant flower or shoot morphology (4 mutants). To whether these mutants were due to transposon insertions, a series of Southern blot experiments was performed on 28 families, comparing in each case several mutant plants with others showing the wild-type phenotype. A preliminary analysis revealed in 4 of the 28 families analyzed a common, novel DsA fragment in all mutant plants, which was present only in heterozygous plants with wt phenotype, as expected for DsA insertion mutations. These four mutants included two showing embryo lethality, one with dark green, unexpanded rosette leaves and stunted inflorescences, and one with curly growth of stems, leaves and siliques. Further evidence for DsA insertion mutations was obtained for one embryo lethal mutant and for the stunted mutant, while in case of the second embryo lethal mutant, the DsA insertion could be separated from the mutant locus by genetic recombination.

Genetic evidence for an essential role of brassinosteroids in plant development

“…Cabbage mutants were isolated in the course of a two-component Ac/Ds transposon mutagenesis as described by Altmann et aL (1995).…”

“…Here we describe the characterization of three extremely dwarfed mutants, cbb l (dwfl-6), cbb2, and cbb3, isolated by transposon mutagenesis in Arabidopsis thaliana (Altmann et aL, 1995) and demonstrate that the cbb mutations affect the biosynthesis and perception of brassinosteroids which are required for the proper control of plant development. cbbl, cbb2, and cbb3 Three mutants named cabbage1, cabbage2, and cabbage3 (cbbl, cbb2, cbb3) exhibiting a strongly stunted axis, a reduced root system, small dark green leaves and a very compact rosette structure ( Figure 1) were isolated from an Arabidopsis mutant collection generated by transposon mutagenesis (Altmann et aL, 1995).…”

“…cbbl, cbb2, and cbb3 Three mutants named cabbage1, cabbage2, and cabbage3 (cbbl, cbb2, cbb3) exhibiting a strongly stunted axis, a reduced root system, small dark green leaves and a very compact rosette structure ( Figure 1) were isolated from an Arabidopsis mutant collection generated by transposon mutagenesis (Altmann et aL, 1995). Soil-grown cbbl mutant plants illuminated with white light showed a maximum height of 6.18 _+ 0.74 cm in comparison with 28.68 _+ 1.67 cm for the wild-type plants, while cbb2 and cbb3 plants only reached a size of less than 1 cm.…”

High-Throughput Screening for Induced Point Mutations

“…Unlike methods that provide only knockout mutations (e.g. Altmann et al, 1995), TILL-ING yields a traditional allelic series of point mutations. This will be especially valuable for essential genes, where sublethal alleles are required for phenotypic analysis.…”

Characterization of the transposition pattern of the Ac element in Arabidopsis thaliana using endonuclease I- Sce I

“…Three out of five mutations were not linked to the transposed element. The occurrence of such unlinked mutations during Ac transposition was previously reported in Arabidopsis (56,57). One possible explanation for this phenomenon has been proposed: the transposed element might have been retransposed from the first transposed site, in which mutations might have been generated to an unlinked chromosomal site.…”