Site-selective and hydrolytic two-strand scission of double-stranded DNA using Ce(IV)/EDTA and pseudo-complementary PNA.
By combining Ce(IV)/EDTA with two pseudo-complementary peptide nucleic acids (pcPNAs), both strands in double-stranded DNA were site-selectively hydrolyzed at the target site. Either plasmid DNA (4361 bp) or its linearized form was used as the substrate. When two pcPNAs invaded into the double-stranded DNA, only the designated portion in each of the two strands was free from Watson-Crick base pairing with the counterpart DNA or the pcPNA. Upon the treatment of this invasion complex with Ce(IV)/EDTA at 37 degrees C and pH 7.0, both of these single-stranded portions were selectively hydrolyzed at the designated site, resulting in the site-selective two-strand scission of the double-stranded DNA. Furthermore, the hydrolytic scission products were successfully connected with foreign double-stranded DNA by using ligase. The potential of these artificial systems for manipulation of huge DNA has been indicated.
Yamamoto
Yoji
Y
Research Center for Advanced Science and Technology, The University of Tokyo, 4-6-1 Komaba, Meguro-ku, Tokyo 153-8904, Japan. komiyama@mkomi.rcast.u-tokyo.ac.jp
Uehara
Akihiko
A
Tomita
Takafumi
T
Komiyama
Makoto
M
D001483:Base Sequence; D002563:Cerium / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D011088:DNA Ligases / Q000378:metabolism; D004278:DNA, Superhelical / Q000378:metabolism; D004492:Edetic Acid / Q000031:analogs & derivatives* / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay; D006868:Hydrolysis; D008969:Molecular Sequence Data; D020135:Peptide Nucleic Acids / Q000737:chemistry*; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000378:metabolism; D013379:Substrate Specificity
D004278:DNA, Superhelical; D020135:Peptide Nucleic Acids; D002563:Cerium; D004247:DNA; D004492:Edetic Acid; D015719:Single-Strand Specific DNA and RNA Endonucleases; D011088:DNA Ligases