A genetic analysis of DNA sequence requirements for Dissociation state I activity in tobacco.

Abstract

Our objective was to test whether the double Ds structure correlated with Dissociation state I activity (i.e., high frequency of chromosome breakage and low frequency of reversion) in maize exhibited similar properties in tobacco. A genetic assay was established to test double Ds and related structures for their ability to cause loss of the linked marker genes streptomycin phosphotransferase and beta-glucuronidase in transgenic tobacco. An engineered double Ds element and a simple Ds element showed behavior consistent with that of state I and state II Ds elements, respectively, as described for maize. DNA structural rearrangements accompanied marker gene loss. Dissection of the double Ds structure showed that a left end and a right end of Ds in direct orientation were sufficient for the instability observed. This result suggested that left and right ends of Ds in direct orientation can participate in aberrant transposition events, consistent with two different models for double Ds-induced chromosome breakage proposed previously. Both models predict that the inversion of a half Ds element accompanies the aberrant transposition event. Such an inversion was detected by polymerase chain reaction experiments in tobacco and maize only when Activator activity was present in the genome.