DIFFERENTIAL METHYLATION AND EXPRESSION OF THE BETA-GLUCURONIDASE AND NEOMYCIN PHOSPHOTRANSFERASE GENES IN TRANSGENIC PLANTS OF POTATO CV BINTJE

Differential methylation and expression of the beta-glucuronidase (GUS) and neomycin phosphotransferase (nptII) genes were studied. Transgenic root clones and regenerated plants of potato cv. Bintje were obtained after transformation with the Agrobacterium tumefaciens strain LBA 1060KG harbouring an agropine wild type Ri plasmid pRi1855 and the binary plasmid pBI121 with the nptII and GUS genes. All seven transgenic root clones and plants regenerated from them were resistant to kanamycin. However, with regard to beta-glucuronidase activity, only four root clones and their regenerants were positive. Though the regenerants of the three GUS negative root clones contained at least four copies of the GUS gene, no GUS mRNA was detected. The suppression of GUS expression was not due to an overall enhancement of methylation of DNA. By using methylation sensitive restriction enzymes, DNA hybridization patterns revealed that the promoter and coding region of the GUS gene was strongly methylated only in the GUS negative shoot lines, irrespective of the number Of genes integrated. In contrast, kanamycin resistance of all shoot lines was consistent with the absence of methylation of the nptII gene. Finally it was found that the reactivation of the GUS gene, occurring during prolonged root culture was accompanied by stable demethylation.