Electrophoresis, Vol.33, No.23, 3429-3436, 2012
Mapping and sequencing DNA using nanopores and nanodetectors
Even prior to the introduction of capillary DNA sequencers, nanopores were discussed as a low-cost, high-throughput substrate for sequencing. Since then, other next-generation sequencing technologies have been developed and achieved widespread use, but nanopores have lagged behind due to difficulties in generating usable sequence data. The practical and theoretical issues of translocation speed and signal detection encountered when attempting to sequence DNA with nanopores are discussed. Various methods that different laboratories have used to overcome difficulties in biologically based and solid-state nanopores are also presented. Different approaches designed to circumvent the overriding issue of detecting signals from individual bases in a time-resolved manner in nanopores are described. For example, genomic positional sequencing utilizes hybridization of short oligonucleotide probes to very long DNA templates and then detects these probes by variations in current blockade in solid-state nanodetectors. The positions of the probes relative to each other and relative to the ends of the DNA are determined by measuring the time between current blockade peaks. By assembling many such measurements, it is possible to overcome the problems encountered when attempting to sequence DNA at high speed in nanopores, providing the potential for true de novo sequencing of large genomes on a routine basis.