EVIDENCE FOR GENOMIC CHANGES IN TRANSGENIC RICE (ORYZA-SATIVA L) RECOVERED FROM PROTOPLASTS

The occurrence of genomic modifications intransgenic rice plants recov ered from protoplasts and their transmission to the self-pollination p rogeny has been verified with the random amplified polymorphic DNA (RA PD) approach. The plant was the Indica-type rice (Oryza sativa L.) cul tivar Chinsurah Bore II. The analysed material was: (1) microspore-der ived embryogenic rice cells grown in suspension culture, (2) transgeni c plants recovered from protoplasts produced from the cultured cells a nd (3) the self-pollination progeny (two successive generations) of th e transgenic plants. DNA purified from samples of these materials was PCR-amplified with different random oligonucleotide primers and the am plification products were analysed by agarose gel electrophoresis. Ban d polymorphism was scored and used in band-sharing analyses to produce a similarity matrix. Relationships among the analysed genomes were ex pressed in a dendrogram. The extensive DNA changes evidenced in cultur ed cells demonstrate the occurrence of somaclonal variation in the mat erial used to produce protoplasts for gene transfer. Quantitatively re duced DNA changes were also found in the resulting transgenic plants a nd in their self-pollination progenies. While confirming the stability of the foreign gene in transgenic plants, this work gives molecular e vidence for the occurrence of stable genomic changes in transgenic pla nts and points to in vitro cell culture as the causative agent. RAPDs are shown to be a convenient tool to detect and estimate the phenomeno n at the molecular level. The methodology is also proposed as a fast t ool to select those transgenic individuals that retain the most balanc ed genomic structure and to control the result of back-crosses planned to restore the original genome.