SOMATIC VARIATION DURING LONG-TERM SUBCULTURING OF PLANT-CELLS CAUSEDBY INSERTION OF A TRANSPOSABLE ELEMENT IN A PHENYLALANINE AMMONIA-LYASE (PAL) GENE

We have identified a new En/Spm-like transposable element, Tdc1, in th e 5' flanking region of a phenylalanine ammonia-lyase gene (gDcPAL1) t hat is normally induced by transferring cells of carrot suspension cul tures to fresh liquid medium (transfer or dilution effect). The initia l integration into gDcPAL1 occurred more than 4 years after culture in itiation. Tdc1 was first detected in gDcPAL1 genomic clones of a genom ic library made from cells of the same cultured cell line 7 years afte r its initiation and thus following repeated subculturing. Twelve year s after initiation, about 5-10% of the cells had Tdc1 inserted into th e gDcPAL1 gene, indicating that Tdc1 insertion into gDcPAL1 occurred i n one (or more) cell(s) during the first 4-7 years of subculturing. Th ese mutant cells did not disappear during numerous passages; instead t he proportion of cells having this Tdc1 inserted into gDcPAL1 has been increasing over the last 5 years. The promoter activity and the induc ibility by transfer/dilution of the gDcPAL1 gene harboring Tdc1 is red uced relative to wild type. Finally, we show that insertion of a trans posable element is one of the mechanisms that can cause variation of p lant cell cultures during repeated subculture.