Stochastic Detection of Terrorist Agents and Biomolecules in a Biological Channel - Nanopores: Sensing and Fundamental Biological Interactions - page 327

sequencing was recently provided by Branton et al. [ 64 ]. One key advantage of
utilizing a nanopore to sequence DNA molecules is that it can read the linear
sequence of nucleotides from the native DNA sample without copying the DNA
and without incorporating labels. Optimal implementation of this method would
include: “sequencing without amplification or modification, and would provide very
long sequence reads (tens of thousands to millions of bases) rapidly and at suffi-
ciently high redundancy to produce assembled sequence of high quality” [ 62 ].
However, as pointed out by Branton and co-workers, one of the major hurdles of
utilizing nanopores to sequence single-stranded DNA (ssDNA) molecules is the
rapid DNA translocation velocity (~1-3 s/base) through the nanopore so that the

accurate detection of single nucleotide bases could not be achieved with the cur-
rently available single-channel recording technique [ 63 ]. In the past decade, various
approaches have been used to slow ssDNA translocation, including immobilization
of DNA polynucleotides with streptavidin [ 65 ], formation of DNA-hemolysin
rotaxane [ 20 ], and formation of double-strand DNA [ 19 ]. Furthermore, single
nucleotide bases could be captured by using a host compound such as

m

-cyclodextrin
[ 66 ]. Using this host-guest interaction approach, single nucleotide base A, T, G, C
were successfully differentiated. In addition, experimental physical conditions such
as temperature, voltage bias, viscosity, and application of an alternating electric field
was also found useful to control the DNA translocation rate [ 67 - 69 ].
Recently, it was reported that the use of ionic liquid solutions as the background
electrolyte instead of the commonly used KCl/NaCl solution increased the values of
the residence time of liquid explosives and monovalent cations in the

b

HL pore
[ 14 ]. The feasibility of utilizing organic salt solutions to slow the translocation of
ssDNA in the

a

HL pore was further investigated [ 70 ]. It was found that in addition
to the rapid DNA translocation events with the mean residence time similar to that
in the NaCl/KCl solution, another type of events with the translocation rates on the
order of hundreds of microseconds per nucleotide base were identified (Fig. 13.10 ).
Since the mean residence time of these long-lived events increased linearly with an
increase in the DNA length [ 70 ], they were suitable for the analysis of the length
and structure of a polynucleotide molecule. More recently, the effect of alkaline pH
on DNA translocation was also systematically investigated by Maglia and co-
workers [ 21 ]. It was reported that the frequency of ssDNA translocation through
the wild-type

a

HL pore decreased with an increase in the pH of the buffer solution.
When pH was larger than 10.7, no events could be identified. Further experiment
showed that ssDNA translocation events were observed in the mutant (M113R) 7
pore even at pH 11.7. These results suggest that the structure of

a

HL pores is stable
enough to tolerate high pH. Furthermore, ssDNA translocation is dependent upon
the charge distribution within the lumen of the

a

HL channel [ 17 ]. The finding is
significant in that the translocation of ssDNA through the nanopore could be
manipulated via introduction of various surface functions into the nanopore, and
double-stranded DNA was able to be studied using the

a

a HL pore approach at high

pH (Fig. 13.11 ).