SELECTION FOR KANAMYCIN RESISTANCE IN TRANSFORMED PETUNIA CELLS LEADSTO THE COAMPLIFICATION OF A LINKED GENE

A cell suspension culture was established from a transgenic petunia (P etunia hybrida L.) plant which carried genes encoding neomycin phospho transferase II(nptII) and beta-glucuronidase (uidA, GUS). Two selectio n experiments were performed to obtain cell lines with increased resis tance to kanamycin. In the first, two independently selected cell line s grown in the presence of 350 mu g/ml kanamycin were eight to ten-fol d more resistant to kanamycin than unselected cells. Increased resista nce was correlated with amplification of the nptII gene and an increas e in nptII mRNA levels. Selection for kanamycin resistance also produc ed amplification of the linked GUS gene, resulting in increased GUS mR NA levels and enzyme activity. Selected cells grown in the absence of kanamycin for twelve growth cycles maintained increased copy numbers o f both genes, and GUS enzyme activity was also stably overexpressed. I n a second selection experiment, a cell line grown continuously in med ium containing 100 mu g/ml kanamycin exhibited higher nptII and GUS ge ne copy numbers and an increase in GUS enzyme activity after eleven gr owth cycles. In this cell line, amplification of the two genes was acc ompanied by DNA rearrangement.