[Ribozyme targeted the point-mutation of activated oncogene inhibits its expression in vivo]

We demonstrated previously that the hammerhead ribozyme designed by us against the activated oncogene T24-ras cleaved the target mRNA specifically and efficiently in vitro. To study its properties in vivo, we cloned the DNA fragment encoding the ribozyme into the eukaryotic expression vector pSMG and transfected it into the T24-ras gene transformed NIH3T3 cell lines. The intracellular cleavage of T24-ras mRNA was evaluated on either the cytological level or the molecular level. The malignant cells expressing ribozyme were partially reversed in morphology as evidenced by slower growth speed, partial recovery of contact-inhibition, reduced frequency of colony forming in soft agar and close-to-normal behaviour in agglutination teat. The primer extension experiment verified that the ribozyme had efficiently cleaved the transcripts of T24-ras gene at the target site in vivo.