Single-molecular AFM probing of specific DNA sequencing using RecA-promoted homologous pairing and strand exchange.
The specific sequence in a linearlized double-stranded DNA target has been identified at a single-molecular level by atomic force microscopy (AFM). This was accomplished using RecA-coated, single-stranded DNA probes which were paired with a specific complementary DNA sequence in a linear double-stranded DNA target by strand-exchange reaction at a homologous sequence site with target DNA. The sites of interaction between the nucleoprotein filaments and the double-stranded DNA targets were directly visualized by AFM in solution containing 4 mM magnesium acetate. Measurements of the position of RecA-coated probes paired to individual target DNA showed that DNA probes specifically paired at their corresponding homologous target sequences. Strand exchange promoted by RecA and the visualization by AFM provided a rapid and efficient way to identify homologous sequence on a single-molecule target DNA.
Seong
G H
GH
Department of Biological Information, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Japan.
Niimi
T
T
Yanagida
Y
Y
Kobatake
E
E
Aizawa
M
M
D001483:Base Sequence; D004247:DNA / Q000737:chemistry*; D015342:DNA Probes; D018625:Microscopy, Atomic Force / Q000379:methods*; D011938:Rec A Recombinases / Q000737:chemistry*
D015342:DNA Probes; D004247:DNA; D011938:Rec A Recombinases