Following up on information from crnano
The Harvard Nanopore Group is working on the development of solid state nanopores and our studies of DNA translocation through these nanopores suggest how a nanopore could be the core of an instrument capable of inexpensive de novo sequencing. They are investigating and developing the basic science and technology required to build a nanopore based instrument that should be able to sequence a mammalian genome for <$1,000 and that meets the following requirements:
a. High-speed sequential identification of the DNA's nucleotides directly on the basis of their distinct physical or electrical properties;
b. Very long, indefinite length reads. Analysis and assembly is a bottle-neck in de novo sequencing and limits re-sequencing when copy number polymorphism or variable indels are to be identified in heterozygous genomes;
c. The requisite sequence coverage (7.7-fold coverage, 6.5-fold coverage in Q20 bases) using genomic DNA from <106 cells with no amplification and minimal preparative steps. Otherwise, amplification or other preparatory steps become limiting.
A biased nanopore in an insulating membrane that separates two ionic solution-filled compartments translocates DNA molecules in sequencial nucleotide order between probes that serve as emitter and collector of a tunneling “microscope.” In response to a voltage bias (labeled “ - ” and “+”) across the membrane, ssDNA molecules (yellow) in the “-” compartment are driven, one at a time, into and through the nanopore. Elevated temperatures and denaturants maintain the DNA in an unstructured, single-stranded form.
Nucleotide bases could be sensed by the currents of a tunneling microscope would be ~ 100,000 – 1,000,000/bases per second if the microscope could follow the molecule's length at that rate and over a sufficient range.
If they are able to resolve each base as it passes through a nanopore at the rate of 10,000 bases/sec, an instrument with an array of 100 such nanopores could produce high-quality draft sequence of one mammalian genome in ~20 hours.