Transactivation of Ds elements in plants of lettuce (Lactuca sativa).
The maize transposable element, Activator (Ac), is being used to develop a transposon mutagenesis system in lettuce, Lactuca sativa. In this paper, we describe somatic and germinal transactivation of Ds by chimeric transposase genes in whole plants. Constructs containing either the Ds element or the Ac transposase open reading frame (ORF) were introduced into lettue. The Ds element was located between either the 35S or the Nos promoter and a chimeric spectinomycin resistance gene (which included a transit peptide), preventing expression of spectinomycin resistance. The genomic coding region of the Ac transposase was expressed from the 35S promoter. Crosses were made between 104 independent R1 plants containing Ds and three independent R1 plants expressing transposase. The excision of Ds in F1 progenies was monitored using a phenotypic assay on spectinomycin-containing medium. Green sectors in one-third of the F1 families indicated transactivation of Ds by the transposase at different developmental stages and at different frequencies in lettuce plants. Excision was confirmed using PCR and by Southern analysis. The lack of green sectors in the majority of F1 families suggest that the majority of T-DNA insertion sites are not conducive to excision. In subsequent experiments, the F1 plants containing both Ds and the transposase were grown to maturity and the F2 seeds screened on medium containing spectinomycin. Somatic excision was again observed in several F2 progeny; however, evidence for germinal excision was observed in only one F2 family.
Yang
C H
CH
Department of Vegetable Crops, University of California, Davis 95616.
Carroll
B
B
Scofield
S
S
Jones
J
J
Michelmore
R
R
D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D017931:DNA Primers; D004251:DNA Transposable Elements; D008969:Molecular Sequence Data; D009713:Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism; D010641:Phenotype; D000198:Spectinomycin / Q000494:pharmacology; D015533:Transcriptional Activation; D019895:Transposases; D014675:Vegetables / Q000235:genetics*
D017931:DNA Primers; D004251:DNA Transposable Elements; D000198:Spectinomycin; D009713:Nucleotidyltransferases; D019895:Transposases