Solid-State Nanopores: Methods of Fabrication and Integration, and Feasibility Issues in DNA Sequencing - Nanopores: Sensing and Fundamental Biological Interactions - page 188

sugar-phosphate backbone of a DNA. It is truly an amazing physical phenomenon
that DNA polymerases, a complex molecular machinery, can resolve individual
bases with high accuracy during the DNA synthesis of a cell cycle.
The DNA sequencing techniques developed by Sanger [ 18 ], Gilbert [ 19 ],
and others take advantage of this unique capability of DNA polymerases. However,
the very process that makes Sanger method possible is also the limiting factor in
reducing the cost while raising the speed of DNA sequencing.

8.4.1 Hybridization-Assisted Nanopore Sequencing (HANS)

It was proposed by Ling et al. [ 14 ] that the solid-state nanopore technology
developed so far may be adequate to implement a low-cost DNA sequencing strategy,
namely, the hybridization assisted nanopore sequencing (HANS). The HANS
approach borrows the idea of a known technique called sequencing-by-hybridization
(SBH) derived from the ideas in “Southern blot” [ 20 ], but HANS differs from SBH
in terms of the way DNA sequences are extracted.
In SBH, small pieces of short oligonucleotides with known sequences are used
as “probes” to hybridize an unknown DNA. From the overlapping parts of hybri-
dizing oligonucleotides, one can in principle reconstruct the sequence of the
unknown DNA using a computer if there are no repetitions of DNA sequences.
However, the genomic DNA is known to have large amount of repeats, preventing
SBH from becoming a viable sequencing technology.
As shown in Fig. 8.9 , a single-stranded ssDNA with unknown sequence is
hybridized with two identical short strands of oligonucleotides and is driven through
a solid-state nanopore. The ionic current through the nanopore is suppressed when
the DNA enters the pore, as usual. The parts of the DNA with matching oligonucleo-
tides strand should suppress the ionic current by a larger magnitude. It is expected

X X X X X X X X X X X X X X X
A A G A T

A A T T C T A A C A T T C T A

A A G A T A A G A T A A G A T

T T A A G

T T G T A

Fig. 8.9 The basic concept of HANS in obtaining the DNA sequence of the ssDNA based on the
positions ( top panel ) of the hybridizing oligonucleotides measured ( lower panel ) by the ionic
current traces for each oligonucleotide [ 14 ]