No transformant was obtained when pCZA168(bla, tsr, Tn5096, ColEI rep. Strep repts) was used to transform S. hygropinocus RF220. pIJ702 isolated from S. hygroscopicus N103 was transformed into RF220 at a low frequency. pIJ702 plasmid was cured in RF220 transformant and it was re-transformed into its cured FR220 strain, but the transformation frequency was not increased significantly, suggesting that restriction-modification system in FR220 was existent and complicated. Four transformants containing pCZA168 were obtained, when the RF220 strain was grown in medium with ampicillin, glycine and the protoplast was stored at -70 degrees C. Restriction analysis of plasmid from transformants indicated that the DNA fragment from E. coli in pCZA168 was deleted. With transposition of Tn5096, two mutants blocked in antibiotic biosynthesis of 120 and some mutants with variation in antibiotic level were obtained, this showed that the Tn5096 transposed in different positions of chromosomal DNA in RF220 and resulted in the production of 120 in different level.