Graphene nanopore-microcavity-solid-state nanopore structure based DNA sequencing device and method

Abstract

The invention discloses a graphene nanopore-microcavity-solid-state nanopore structure based DNA sequencing device and a method. The sequencing device is mainly composed of a graphene nanopore-equipped graphene microstrip, an inverted pyramid-shaped microcavity in a silicon-based substrate, and a solid-state nanopore at the top of the microcavity. During sequencing, a sequencing reaction chamber is divided into two parts by the graphene nanopore-microcavity-solid-state nanopore structure. A single-stranded DNA molecule passes through the graphene nanopore linearly under the effect of an electrostatic field, then enters the inverted pyramid-shaped microcavity, and finally comes out from the solid-state nanopore. Weak current measuring equipment is utilized to measure longitudinal ionic current blocking caused by the DNA molecule passing activity in the nanopore and the transverse conductance change around the nanopore in the graphene microstrip. Further, a synchronous data recording and processing system is employed for analytical calculation of bi-directional data, thus realizing sequence analysis of the single-stranded DNA molecule.

Description

technical field [0001] The invention belongs to the technical field of DNA molecular sequencing, in particular to a DNA sequencing device and method based on a graphene nanopore-microcavity-solid nanopore structure. Background technique [0002] Deoxyribonucleic acid (DNA) sequencing technology is one of the core technologies of modern life science research. In order to achieve the goals of thousand-dollar human genome (TDG) and hundred-dollar human genome (HDG), and promote individualized disease diagnosis and treatment, a low-cost, high-throughput direct sequencing method is urgently needed. Nanopore-based single-molecule sequencing is considered to be the most promising key technology to achieve the above goals. [0003] So far, among the various nanopore-based DNA single-molecule sequencing methods that have been reported, the ion current blocking method is the earliest and the most widely studied. The basic principle of this method is as follows. The sequencing reacti...

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Problems solved by technology

[0004] However, this nanopore ionic current blocking method faces some fundamental problems for practical application.

The biomolecular nanopores used in the early stage (typically represented by nanopores composed of protein α-hemolysin) have poor stability, short lifespan, and are extremely sensitive to environmental factors, and the pore size of biomolecular nanopores is difficult to artificially Control, the internal pore size is only about 1.5nm, not suitable for the detection of different nucleic acid molecules

Although the solid-state nanopore (Solid-state nanopore) commonly used at present overcomes the above-mentioned shortcomings of biomolecular nanopores, it also has the following problems: First, the channel length of the solid-state nanopore is usually more than 5 nm, and can accommodate more than ten bases. This size is too long for sequencing to resolve the current change caused by a single base; secondly, when a single nucleotide occupies a nanopore, only about 100 ions pass through the nanopore, and 4 bases have only a few ions in the structure. The difference of each atom, the change of ionic current caused by this subtle structural difference is very weak, so that it is difficult for researchers to distinguish each base; third, the sequencing method based on solid-state nanopore cannot effectively control DNA The speed of passing through the nanopore is too fast, resulting in a low recognition rate of base detection

[0005] Various nanopore sequencing methods with the help of other auxiliary means, such as fluorescent label-assisted nanopore sequencing, hybridization-assisted nanopore sequencing, tunneling current-assisted nanopore sequencing, and probe-modified nanopore sequencing, are essentially It is still an indirect sequencing method, and there are problems such as complex equipment, low speed, and high cost

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