Sequence-dependent hydrolysis of RNA using modified oligonucleotide splints and RNase H

We found that, in the presence of chimeric oligonucleotides containing complementary deoxyribo- and 2'-O-methylnucleosides, a nonaribonucleotide, [5'-32P]pACUUACCUG, was cleaved specifically upon treatment with RNase H. When 3'm(UG)d(AATG)m(GAC)5' was used as a hybridization strand, pACUUACCUG was cleaved between C6 and C7 to yield pACUUAC. In the presence of 3'm(UGAA)d(TGGA)m(C)5', the nonaribonucleotide was hydrolyzed, mainly between U8 and C9, to give pACUUACCU. This method will have a variety of applications in the field of RNA engineering.