In vivo footprinting experiments have been used to analyze the binding of trans-acting regulatory factors in the 5' flanking region upstream of the alcohol dehydrogenase (Adh) gene from Arabidopsis thaliana. Protein-DNA interactions were detected by dimethyl sulfate footprinting and genomic sequencing, using an A. thaliana cell suspension culture that constitutively expressed the Adh gene. Several distinct footprinting domains have been characterized, and the potential effects of the corresponding trans-acting factors have been inferred from a comparison with data from the maize alcohol dehydrogenase-1 (Adh1) gene. One binding site is similar in sequence to one of the anaerobic response elements (ARE) of the maize gene, which has also been shown to bind to a trans-acting factor. Several of the remaining binding sites apparently represent a class of elements sharing the sequence 5'-GTGG-3' within their footprint.Comparisons with maize Adh1 in vivo protein interactions reveal that the elements of Adh promoter structure are highly conserved, but the relative and absolute positions of the elements are variable.

S1 nuclease was used to probe the architectural characteristics of the maize alcohol dehydrogenase-1 gene promoter. Three sites were identified as hypersensitive to S1 digestion in supercoiled, but not linear plasmids containing the Adh1 gene. The sites mapped to areas located 65, 330 and 800 base pairs 5' to the start of transcription. In each case, the strand specific nicking pattern was determined with nucleotide level precision. The -65 site was found to be a homopurine/homopyrimidine tract. The -330 site mapped to the boundaries of a region of high Z-DNA potential and the -800 site mapped to a non-descript sequence. The possible biological significance of these sites is discussed.

Transgenic plants of lettuce cultivar (cv.) 'South Bay' were produced by using Agrobacterium tumefaciens vectors containing the /3-glucuronidase (GUS) reporter gene and the NPT II gene for kanamycin resistance as a selectable marker. High frequency of transformation, based on kanamycin resistance and assays for GUS expression, was obtained with 24 to 72-h-old cotyledon explants cocultivated for 48 h with Agrobacterium tumefaciens. After the cocultivation period, the explants were placed in selection medium containing 50 or 100mgl -I of kanamycin, 100mg1-1 cefotaxime and 500mg1-1 carbenicillin for 10 days. Surviving explants were transferred every 14 days on shoot elongation medium. Progenies of R 0 plants demonstrated linked monogenic segregation for kanamycin resistance and GUS activity.