Development of polymer latex particles for selective cleavage of mismatched DNA and their application for DNA diagnosis.
We developed functional polymer latex particles that can catch and cleave mismatched DNA selectively and propose a new mismatch detection system using the functional particles. The aimed particles possess two functional units composed of mismatch binding protein (MutS) and an anthraquinone derivative (AQ), a light-activated agent that photocleaves dsDNA. Use of the functional particles made it possible to discriminate complementary and mismatched DNAs and photocleave mismatched DNA selectively. The efficiency of photocleavage of mismatched DNA by the functional particles increased with UV irradiation time. It was also found that the functional particles were reusable and had dissociation constants (K(d)) of 1000 and 68.5 nM for G/C homoduplex and G/T heteroduplex, respectively. Using the functional particles and a dsDNA-binding fluorescent dye, SYBR-Gold, we could construct the system for detection of mismatched DNA that was 40 base pairs. The functional particles prepared in this study will be an absolutely new tool for mismatch detection in DNA diagnosis.
Oba
Shinsuke
S
Department of Applied Chemistry, Faculty of Science and Technology, Keio University, Kouhoku-ku, Yokohama 223-8522, Japan.
Hatakeyama
Mamoru
M
Handa
Hiroshi
H
Kawaguchi
Haruma
H
D000251:Adenosine Triphosphatases / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism; D020137:Base Pair Mismatch / Q000235:genetics*; D001483:Base Sequence; D004247:DNA / Q000235:genetics* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism; D004800:Enzymes, Immobilized / Q000378:metabolism; D005820:Genetic Testing / Q000379:methods*; D007840:Latex / Q000737:chemistry*; D046529:Microscopy, Electron, Transmission; D008863:Microspheres; D051719:MutS DNA Mismatch-Binding Protein; D010316:Particle Size; D010782:Photolysis; D013050:Spectrometry, Fluorescence; D014105:Tosyl Compounds / Q000737:chemistry
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D004800:Enzymes, Immobilized; D007840:Latex; D014105:Tosyl Compounds; D004247:DNA; D000251:Adenosine Triphosphatases; D051719:MutS DNA Mismatch-Binding Protein