A double knockout mutant of acyl-glucose-dependent anthocyanin glucosyltransferase genes in Delphinium grandiflorum.

Blue coloration in delphinium flowers arises from 7-polyacylated anthocyanins which are modified alternately with acyl and glucosyl residues at the 7 position of the aglycone. Previously, we identified two independent genes for acyl-glucose-dependent anthocyanin 7-(6-(p-hydroxybenzoyl)-glucoside) glucosyltransferases (AA7BG-GT); recombinant proteins from the two cDNAs were produced in Escherichia coli and showed AA7BG-GT activity in vitro. Here, a double knockout mutant of both genes was found to lack modification of the second glucosyl residue following further acyl and glucosyl modifications. Both genes in the double mutant had nucleotide sequence changes and deletions that disrupted their transcripts and caused loss of AA7BG-GT activity in sepals. These results provide genetic confirmation that both genes are responsible for AA7BG-GT enzyme activity.