Identification of hairy root loci in the T-regions of Agrobacterium rhizogenes Ri plasmids

Summary Agrobacterium rhizogenes induces root formation at the wound site of inoculation in plants and inserts a fragment of its plasmid (Ri) into the plant nuclear DNA. Parts of the transferred region (T-region) of the Ri plasmid of A. rhizogenes strain A4 or 8196 are cloned in Escherichia coli. Insertions of the E. coli lacZ coding region into the hybrid plasmids were made in vivo using transduction by miniMu. Twenty insertions localized in the T_L-DNA of pRiA4 (or pRi1855) and 2 inserts in the T-DNA of pRi8196 were obtained in E. coli. One of the T_L-DNA insertions is saved up because it is linked to an internal T-DNA deletion; the others because they confer a lactose plus phenotype on E. coli; this indicates that the T-DNA harbours sequences that are expressed in E. coli. Fifteen of these T-DNA insertions were transfered to Agrobacterium where they substitute the corresponding wild-type T-DNA of the Ri plasmid by homologous recombination. These strains corresponding to insertion-directed mutagenesis were used to inoculate Daucus carota slices and stems and leaves of Kalanchoe daigremontiana. The two insertions strains obtained in the T-DNA of pRi8196 are avirulent on K. daigremontiana; but their phenotypes differ on D. carota slices, suggesting that insertions affect distinct loci on the T-DNA involved in hairy root formation. Only one insertion out of the twenty obtained in the T_L-DNA of pRiA4 (or 1855) induces a loss of virulence on leaves of K. daigremontiana. However the T_L-DNA deletion harbouring strain induces a loss of virulence on D. carota and K. daigremontiana (stems and leaves), confirming the importance of the T_L-DNA for hairy root induction. re]19850711 rv]19851230 ac]19860114