An analysis of mismatched duplex DNA unzipping through a bacterial nanopore.
A 50-base Guide strand was synthesized that consisted of a central 10-base probe sequence flanked by two tracts of 20 adenine residues. Target sequences of 10 bases containing up to three mismatches were prepared and hybridized to the Guide strand in 1 M KCl. The transport of these constructs through single alpha-hemolysin pores was analysed by measuring the current blockade as a function of time. Complementary dsDNA takes significantly longer (840 +/- 60 micro s) to pass through the pore than a sequence of the same length containing a single (590 +/- 45 micro s) and a double (270 +/- 50 micro s) mismatch. Constructs involving three mismatches were indistinguishable from Guide ssDNA transport (120 +/- 30 micro s). The results suggest that dsDNA must unzip as it is transported through the nanopore. Duplexes containing mismatches unzip more quickly and can be distinguished from those with perfect complementarity.
Sutherland
Todd C
TC
Department of Chemistry, University of Saskatchewan, 110 Science Place, Saskatoon, S7N 5C9, Canada.
Dinsmore
Michael J
MJ
Kraatz
Heinz-Bernhard
HB
Lee
Jeremy S
JS
D000225:Adenine / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D020029:Base Pairing; D001692:Biological Transport; D004277:DNA, Single-Stranded / Q000737:chemistry*; D006460:Hemolysin Proteins / Q000737:chemistry*; D008051:Lipid Bilayers / Q000737:chemistry*; D018272:Porins / Q000737:chemistry; D011189:Potassium Chloride / Q000737:chemistry
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D006460:Hemolysin Proteins; D008051:Lipid Bilayers; D018272:Porins; D011189:Potassium Chloride; D000225:Adenine