Controlled Microwave Heating Accelerates Rolling Circle Amplification.
Rolling circle amplification (RCA) generates single-stranded DNAs or RNA, and the diverse applications of this isothermal technique range from the sensitive detection of nucleic acids to analysis of single nucleotide polymorphisms. Microwave chemistry is widely applied to increase reaction rate as well as product yield and purity. The objectives of the present research were to apply microwave heating to RCA and indicate factors that contribute to the microwave selective heating effect. The microwave reaction temperature was strictly controlled using a microwave applicator optimized for enzymatic-scale reactions. Here, we showed that microwave-assisted RCA reactions catalyzed by either of the four thermostable DNA polymerases were accelerated over 4-folds compared with conventional RCA. Furthermore, the temperatures of the individual buffer components were specifically influenced by microwave heating. We concluded that microwave heating accelerated isothermal RCA of DNA because of the differential heating mechanisms of microwaves on the temperatures of reaction components, although the overall reaction temperatures were the same.
Department of Applied Biological Science, Tokyo University of Science, 264 Yamazaki, Noda, Chiba 305-8506, Japan.
Takeo
T
Yoshimura
Department of Applied Biological Science, Tokyo University of Science, 264 Yamazaki, Noda, Chiba 305-8506, Japan.
Takamasa
T
Suzuki
Department of Applied Biological Science, Tokyo University of Science, 264 Yamazaki, Noda, Chiba 305-8506, Japan.
Shigeru
S
Mineki
Graduate School of Life Science and System Engineering, Kyushu Institute of Technology, 680-4 Kawazu, Iizuka, Fukuoka 820-8502, Japan.
Shokichi
S
Ohuchi
D004270:DNA, Circular / Q000235:genetics / Q000528:radiation effects; D004277:DNA, Single-Stranded / Q000235:genetics* / Q000528:radiation effects; D004259:DNA-Directed DNA Polymerase / Q000235:genetics* / Q000528:radiation effects; D008872:Microwaves; D021141:Nucleic Acid Amplification Techniques; D012313:RNA / Q000528:radiation effects
D004270:DNA, Circular; D004277:DNA, Single-Stranded; D012313:RNA; D004259:DNA-Directed DNA Polymerase