Characterization of the maize transposable element Ac by internal deletions

Abstract

We have used the ability of Ac to transpose in tobacco to determine which Ac sequences are required for transposition, using a phenotypic assay for Ac excision from an NPTII gene in which excisions of Ac result in calli resistant to the antibiotic kanamycin (Baker et al., 1987). Here we show that deletion of the Ac DNA which encodes the untranslated leader of the Ac transcript does not prevent Ac excision. A deletion which removes 110 bp including the first 75 bp of long open reading frame prevents Ac excision in tobacco cells. However, it will excise in tobacco cells previously transformed with Ac indicating that deletion of the region prevents the synthesis of a product required for Ac excision. Deletion of the Ac sequences between bp 44 and bp 92 or from bp 75 to bp 181 abolishes, or strongly reduces, transposition in cells which are already transgenic for an active Ac element. This indicates that these deleted elements have lost sequences which are required for the transposon to respond to the transposase, when the enzyme is produced in trans. We also describe a tobacco strain transformed with a Ds element stably inserted wtihin an NPTII gene. This strain is Km and was retransformed with a construct containing the open reading frame (ORF) of the 3.5-kb Ac transcript expressed from a plant promoter. Expression of the cDNA construct promotes excision of the Ds element. These data suggest that the 3.5-kb transcript of Ac encodes the only Ac product required for transposition, i.e. the transposase function.