To evaluate the prospects for transposon mutagenesis in the autogamous
diploid legume Lotus japonicus, the behaviour of the maize transposab
le element Ac was analysed in the progeny;of 38 independent transgenic
plants. The conditions for monitoring donor site excision using histo
chemical localization of beta-glucuronidase activity or the alternativ
e spectinomycin resistance assay were established, and used to follow
Ac mobility through two generations. Somatic excision was monitored as
variegated cotyledons in the T-2 generation and germinal excision eve
nts were scored in segregating T-3 families as complete beta-glucuroni
dase-mediated staining of cotyledons or as a fully green spectinomycin
-resistant phenotype. Using these assays an average germinal excision
frequency of 12% was estimated in the T-3 offspring from variegated pl
ants. The fidelity of the excision assays was ascertained by comparing
the frequency of germinal excision to the frequency of Ac reinsertion
at new positions of the genome. Transposition of Ac in 42% of the pla
nts and detection of the characteristic Ac insertion/excision footprin
ts suggests that insertion mutagenesis with the autonomous maize Activ
ator element is feasible in Lotus japonicus. Parameters influencing Ac
behaviour, such as dosage, position effects and modification of the e
lement itself, were also investigated comparing homozygous and hemizyg
ous plants from the same family and by analysing different transforman
t.