U.S. patent application number 17/370841 was filed with the patent office on 2021-12-30 for nanoscale etching of light absorbing materials using light and an electron donor solvent.
The applicant listed for this patent is Northeastern University. Invention is credited to Meni Wanunu, Hirohito Yamazaki.
Application Number | 20210405533 17/370841 |
Document ID | / |
Family ID | 1000005839398 |
Filed Date | 2021-12-30 |
United States Patent
Application |
20210405533 |
Kind Code |
A1 |
Wanunu; Meni ; et
al. |
December 30, 2021 |
Nanoscale Etching of Light Absorbing Materials using Light and an
Electron Donor Solvent
Abstract
A method for etching a light absorbing material permits directly
writing a pattern of etching of silicon nitride and other light
absorbing materials, without the need of a lithographic mask, and
allows the creation of etched features of less than one micron in
size. The method can be used for etching deposited silicon nitride
films, freestanding silicon nitride membranes, and other light
absorbing materials, with control over the thickness achieved by
optical feedback. The etching is promoted by solvents including
electron donor species, such as chloride ions. The method provides
the ability to etch silicon nitride and other light absorbing
materials, with fine spatial and etch rate control, in mild
conditions, including in a biocompatible environment. The method
can be used to create nanopores and nanopore arrays.
Inventors: |
Wanunu; Meni; (Needham,
MA) ; Yamazaki; Hirohito; (Brighton, MA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Northeastern University |
Boston |
MA |
US |
|
|
Family ID: |
1000005839398 |
Appl. No.: |
17/370841 |
Filed: |
July 8, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
|
16368616 |
Mar 28, 2019 |
11073764 |
|
|
17370841 |
|
|
|
|
62650023 |
Mar 29, 2018 |
|
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C04B 41/5353 20130101;
G03F 7/2053 20130101; G03F 7/0041 20130101; G01N 21/59 20130101;
G03F 7/11 20130101; G03F 7/2043 20130101 |
International
Class: |
G03F 7/20 20060101
G03F007/20; C04B 41/53 20060101 C04B041/53; G01N 21/59 20060101
G01N021/59; G03F 7/11 20060101 G03F007/11; G03F 7/004 20060101
G03F007/004 |
Claims
1. An apparatus comprising a light absorbing material, the
apparatus comprising: an etched feature comprising a nanopore
through the light absorbing material, the nanopore having a
diameter greater than about 1 nanometer and less than about 1
micron; wherein the nanopore has a greater diameter on one side of
the light absorbing material than on an opposite side of the light
absorbing material.
2. The apparatus of claim 1, wherein the light absorbing material
comprises a thickness less than about 1 micron at the location of
the nanopore.
3. The apparatus of claim 1, comprising an array of a plurality of
nanopores.
Description
RELATED APPLICATION
[0001] This application is a divisional of U.S. application Ser.
No. 16/368,616, filed on Mar. 28, 2019, which claims the benefit of
U.S. Provisional Application No. 62/650,023, filed on Mar. 29,
2018. The entire teachings of the above applications are
incorporated herein by reference.
INCORPORATION BY REFERENCE OF MATERIAL IN ASCII TEXT FILE
[0002] This application incorporates by reference the Sequence
Listing contained in the following ASCII text file:
[0003] a) File name: 52002209004_SEQUENCELISTING.txt; created Sep.
17, 2021, 1,070 Bytes in size.
BACKGROUND
[0004] Silicon nitride is a widely used material as a component of
solid-state devices for applications in biotechnology, electronics,
photonics, and other areas. Processing silicon nitride films in
order to provide defined topographical features is a major part of
the silicon industry, and has been for the past few decades a
routine practice for micro- and nanostructure fabrication.
Lithography in silicon nitride typically requires aggressive dry or
wet chemical treatment, which can be accomplished using a
photolithography or electron beam mask layer followed by chemical
processing.
[0005] There is, however, an ongoing need to provide improved
methods of etching silicon nitride and other light absorbing
materials, so that the rate of etching and the spatial extent of
the etching can be more finely controlled, and so that the etching
can be performed with less expense and in milder conditions.
[0006] In addition, there is an ongoing need for methods to shape
materials at the nanoscale. These have received much attention in
recent years, largely due to the need for technologies that allow
the investigation of quantum phenomena at the nanoscale, but also
as vehicles for interfacing with the molecular/biomolecular world.
Nanopore sensors are prime examples of nanoscale devices that sense
small molecules, biomolecules, and their complexes. Sensing in
nanopores is most frequently performed by applying a voltage across
a nanopore that contains two electrolyte baths across it and
measuring as a function of time the flux of ions that cross the
pore. When an analyte (e.g., DNA, protein, or a small molecule)
occupies the pore, the ion flux temporarily changes, affording
indirect electrical detection of biomolecules in a label-free
manner.
[0007] An attractive form of nanopores for future integration into
sensing devices is a silicon chip that contains a freestanding
membrane composed of silicon nitride (SiN.sub.x) or another
material, through which a hole is fabricated using one of various
methods. Nanopore sensors are not "one-size-fits-all"; their
dimension requirements are typically commensurate with the analyte
size. Therefore, the nanopore physical properties, which include
its diameter, length, shape, and interfacial properties, typically
need tailoring for optimal sensing.
[0008] A variety of approaches have been developed for nanopore
fabrication, but there is an ongoing need for techniques that
confer high resolution for biopolymer analysis, for example, for
creating high-resolution pores that are comparable in geometry to
protein-based nanopores used in DNA sequencing.
SUMMARY
[0009] In accordance with an embodiment of the invention, a method
for etching a light absorbing material permits directly writing a
pattern of etching of silicon nitride and other light absorbing
materials, without the need of a lithographic mask, and allows the
creation of etched features of less than one micron in size. The
method can, for example, be used for etching deposited silicon
nitride films, freestanding silicon nitride membranes, and other
light absorbing materials, with control over the thickness achieved
by optical feedback. The etching is promoted by solvents including
electron donor species, such as chloride ions. The method provides
the ability to etch silicon nitride and other light absorbing
materials, with fine spatial and etch rate control, in mild
conditions, including in a biocompatible environment. The method
can be used to create nanopores and nanopore arrays.
[0010] In accordance with one embodiment, there is provided a
method for etching a light absorbing material. The method comprises
contacting the light absorbing material with a solvent comprising
an electron donor species, and exposing an interface between the
solvent and the light absorbing material to light having a
wavelength within an absorption band of a light absorbance spectrum
of the light absorbing material to induce etching of the light
absorbing material, at the interface, by the light and the electron
donor species to create an etched feature of less than 1 micron in
size.
[0011] In further, related embodiments, the light absorbing
material may comprise silicon; and may comprise at least one of a
silicon-containing nitride and a silicon-containing carbide. The
light absorbing material may comprise a chemical formula SiN.sub.x,
where x is greater than or equal to 0 and less than or equal to 2.
The electron donor species may comprise at least one of a halogen
ion and a hydroxide ion; and may comprise a chloride ion. The
method may comprise exposing the interface between the solvent and
the light absorbing material to the light in the absence of a
lithographic mask. The etching may comprise forming a nanopore
through the light absorbing material, the nanopore having a
diameter greater than about 1 nanometer and less than about 1
micron. The light absorbing material may comprise a thickness less
than about 1 micron at the location of the nanopore.
[0012] In other, related embodiments, forming the nanopore may
comprise limiting a power density of the light after a current
measured through the light absorbing material rapidly increases
above a current threshold during the etching. The method may
comprise limiting the current threshold to determine a size of the
nanopore formed by the etching. The etched feature may define a
nanopore having a greater diameter on one side of the light
absorbing material than on an opposite side of the light absorbing
material. The method may further comprise using the etching to form
an array of a plurality of nanopores. The method may further
comprise assisting the etching by applying a voltage across the
light absorbing material to promote dielectric breakdown of the
light absorbing material. The etched feature may be formed
underneath a surface of a structure of which the light absorbing
material forms at least a portion.
[0013] In further, related embodiments, the method may comprise
controlling a thickness of the etched feature using feedback from
an optical measurement of the etched feature. The solvent may be
biocompatible, and the etching may be performed in a biocompatible
environment. The light may have a wavelength between about 10 nm
and about 400 nm; and may have a wavelength between about 400 nm
and about 700 nm. The light may have an average power density of
greater than about 10.sup.5 watts per square centimeter, such as
between about 10.sup.5 watts per square centimeter and about
10.sup.8 watts per square centimeter.
[0014] In other, related embodiments, the method may further
comprise performing a three-dimensional, layer by layer etching of
the light absorbing material. The method may comprise using a light
image projection device to provide the light, to perform the
three-dimensional, layer by layer etching, based on an electrical
signal comprising a three-dimensional etching pattern having
features with dimensions less than 1 micron in size. The etching
may be used to polish a surface of the light absorbing material.
The method may comprise reflecting a beam of the light from a
spatial light modulator onto the light absorbing material.
Reflecting the beam of the light from the spatial light modulator
onto the light absorbing material may comprise reflecting the beam
of the light from a digital micromirror device onto the light
absorbing material. The method may comprise directly controlling a
location of the etched feature on the light absorbing material
based on a location of the light on the light absorbing
material.
[0015] In another embodiment, there is provided an apparatus
comprising a light absorbing material, the apparatus comprising an
etched feature comprising a nanopore through the light absorbing
material, the nanopore having a diameter greater than about 1
nanometer and less than about 1 micron; wherein the nanopore has a
greater diameter on one side of the light absorbing material than
on an opposite side of the light absorbing material.
[0016] In further, related embodiments, the light absorbing
material may comprise a thickness less than about 1 micron at the
location of the nanopore. The apparatus may comprise an array of a
plurality of nanopores.
BRIEF DESCRIPTION OF THE DRAWINGS
[0017] The foregoing will be apparent from the following more
particular description of example embodiments, as illustrated in
the accompanying drawings in which like reference characters refer
to the same parts throughout the different views. The drawings are
not necessarily to scale, emphasis instead being placed upon
illustrating embodiments.
[0018] FIG. 1 is a schematic diagram illustrating a method for
etching a light absorbing material, in accordance with an
embodiment of the invention.
[0019] FIG. 2 is a graph illustrating a light absorbance spectrum
of a light absorbing material, which can be etched by a method in
accordance with an embodiment of the invention.
[0020] FIG. 3 is a schematic diagram illustrating a proposed
mechanism of etching of SiN.sub.x, assisted by chloride ions, in
accordance with an embodiment of the invention.
[0021] FIG. 4 is a schematic diagram illustrating a method for
three-dimensional etching, in accordance with an embodiment of the
invention.
[0022] FIG. 5 is a schematic diagram illustrating use of a method
of etching in accordance with an embodiment of the invention to
polish a surface of a light absorbing material.
[0023] FIG. 6 is a schematic diagram illustrating components used
to control formation of a nanopore, in accordance with an
embodiment of the invention.
[0024] FIG. 7 is a graph illustrating measurements during a method
of limiting the power density of the light after a current measured
through the light absorbing material rapidly increases above a
current threshold during the etching, in accordance with an
embodiment of the invention.
[0025] FIG. 8 is a schematic diagram of a nanopore, having a
greater diameter on a -cis side of a nanopore, as compared with a
-trans side, formed in accordance with an embodiment of the
invention.
[0026] FIGS. 9A-9C are diagrams illustrating controlled
photothermal etching, in accordance with an embodiment of the
invention.
[0027] FIG. 9A is a continuous current trace of a SiN.sub.x and
HfO.sub.2-coated SiN.sub.x pore at T.sub.pore=60.degree. C. for 10
min (V=100 mV) under 532 nm laser irradiation (laser turned on
after 5 s), in an experiment in accordance with an embodiment of
the invention.
[0028] FIG. 9B is a TEM image of 75 nm-thick SiN.sub.x membrane
after 47 mW laser illumination for 10 min in water, 0.4 M sodium
acetate, and 0.4 M KCl (T=300.degree. C.), in an experiment in
accordance with an embodiment of the invention.
[0029] FIG. 9C is a TEM image of a 75 nm-thick SiN.sub.x membrane
after 1.5 mW laser illumination for 8 h in 4 M KCl, in an
experiment in accordance with an embodiment of the invention.
[0030] FIG. 10A is a schematic diagram of photochemical etching on
freestanding SiN.sub.x membrane, in an experiment in accordance
with an embodiment of the invention.
[0031] FIG. 10B shows AFM-measured etch depth profiles, laser at
different laser powers and times, for 75 nm-thick SiNx supported by
2.5-.mu.m-thick silicon oxide on silicon, in an experiment in
accordance with an embodiment of the invention.
[0032] FIG. 11 is a continuous current trace of 75 nm-thick
SiN.sub.x membrane etching in 0.4 M KCl and 4 M KCl using a 10 mV
probing voltage (laser power was 47 mW), in an experiment in
accordance with an embodiment of the invention.
[0033] FIGS. 12A and 12B are diagrams illustrating laser-assisted
dielectric breakdown for nanopore fabrication, in experiments in
accordance with an embodiment of the invention.
[0034] FIG. 12A shows a continuous current trace obtained during
SiN.sub.x laser assisted dielectric breakdown (35 mW laser power,
V=1 V, laser turned on at 0 s), in an experiment in accordance with
an embodiment of the invention.
[0035] FIG. 12B is a scatter plot of pore thicknesses vs breakdown
times for 29 pores with three different laser powers used (P=47 mW
shown in red, P=35 mW shown in blue, and P=24 mW shown in green),
in an experiment in accordance with an embodiment of the
invention.
[0036] FIG. 12C is a diagram showing typical .DELTA.I/I0 vs log
t.sub.d scatter plots obtained for 2.5 kbp DNA translocation
dynamics through a 3.3 nm diameter pore with 2.8 nm effective
thickness at 300 mV from cis (left) and trans (right) side, in an
experiment in accordance with an embodiment of the invention.
[0037] FIGS. 13A and 13B show discrimination of 250 bp and 2.5 kbp
DNA, in experiments in accordance with an embodiment of the
invention.
[0038] FIG. 13A shows continuous 1 s trace excerpts at 200, 250,
and 300 mV for a 2.7 nm pore with 1.7 nm effective thickness for a
mixture of 250 bp (30 nM) and 2.5 kbp (3 nM) dsDNA added to the cis
chamber, in an experiment in accordance with an embodiment of the
invention.
[0039] FIG. 13B shows .DELTA.I/I0 vs log t.sub.d of a mixture of
250 bp and 2.5 kbp at 200, 250, and 300 mV, showing an optimal
resolution between the two lengths at 300 mV, in an experiment in
accordance with an embodiment of the invention.
[0040] FIGS. 14A-14D are diagrams showing current discrimination of
dC and dA homopolymers using an ultrathin laser-assisted breakdown
pore, in experiments in accordance with an embodiment of the
invention.
[0041] FIG. 14A is a schematic diagram of sequentially unzipping
dsDNA to expose a predetermined temporal sequence of homopolymers
to the pore, in an experiment in accordance with an embodiment of
the invention.
[0042] FIG. 14B is a current trace of DNA unzipping in a 1.4 nm
diameter pore with 1.8 nm effective thickness at 200 mV (trace was
low-pass-filtered at 1 kHz), in an experiment in accordance with an
embodiment of the invention.
[0043] FIG. 14C shows I levels of poly(dC) (blue markers) and
poly(dA) (red markers) and .DELTA.I dA-dC (black squares), in an
experiment in accordance with an embodiment of the invention.
[0044] FIG. 14D is a histogram of differential .DELTA.I dA-dC with
the Gaussian fit, in an experiment in accordance with an embodiment
of the invention.
DETAILED DESCRIPTION
[0045] A description of example embodiments follows.
[0046] In accordance with an embodiment of the invention, a method
for etching a light absorbing material permits directly writing a
pattern of etching of silicon nitride and other light absorbing
materials, without the need of a lithographic mask, and allows the
creation of etched features of less than one micron in size. The
method can, for example, be used for etching deposited silicon
nitride films, freestanding silicon nitride membranes, and other
light absorbing materials, with control over the thickness achieved
by optical feedback. The etching is promoted by solvents including
electron donor species, such as chloride ions. The method provides
the ability to etch silicon nitride and other light absorbing
materials, with fine spatial and etch rate control, in mild
conditions, including in a biocompatible environment. The method
can be used to create nanopores and nanopore arrays.
[0047] Sculpting solid-state materials at the nanoscale is an
important step in the manufacturing of numerous types of sensor
devices, such as solid-state nanopore sensors. An embodiment
according to the invention provides laser-induced thinning of
low-stress silicon nitride (SiN.sub.x) membranes and films, and
other light absorbing materials. The light absorbing material, such
as SiN.sub.x, can be etched under exposure to light of an average
intensity on the order of 10.sup.7 W/cm.sup.2, or other intensities
taught herein, with etch rates that are influenced by the
supporting electrolyte. Combining this controlled etching with
dielectric breakdown, an electrokinetic process for making pores,
including nanopores of arbitrary dimensions as small as 1-2 nm in
diameter and thickness, is provided. Refined control over pore
dimensions can expand the range of applications of solid-state
nanopores, for example, for biopolymer sequencing and detection of
specific biomarkers.
[0048] FIG. 1 is a schematic diagram illustrating a method for
etching a light absorbing material, in accordance with an
embodiment of the invention. The method includes contacting a light
absorbing material 102, such as silicon nitride (SiN.sub.x), with a
solvent comprising an electron donor species 104, such as chloride
ions. The interface 106 between the solvent and the light absorbing
material is exposed to light 108 having a wavelength within an
absorption band 210a/b (see FIG. 2) of a light absorbance spectrum
of the light absorbing material 102. The light 108 and the electron
donor species 104 together induce etching of the light absorbing
material 102, at the interface 106, to create an etched feature 112
of less than 1 micron in size.
[0049] FIG. 2 is a graph illustrating a light absorbance spectrum
of a light absorbing material, which can be etched by a method in
accordance with an embodiment of the invention. In this example,
the light absorbing material is silicon nitride (SiN.sub.x), and
two light absorption bands 210a/b are shown; one, 210a, in the
ultraviolet portion of the light absorbance spectrum, and one,
210b, in the visible portion of the light absorbance spectrum. As
used herein, a "light absorption band" is a range of wavelengths in
the light absorbance spectrum of a light absorbing material that
absorbs light at a non-zero absorbance that is sufficient to
promote etching of the light absorbing material by methods taught
herein.
[0050] In accordance with embodiments of the invention, the light
used can, for example, have an ultraviolet wavelength, such as
between about 10 nm and about 400 nm, and can have a visible
wavelength, such as between about 400 nm and about 700 nm. The
light can have an average power density of greater than about
10.sup.5 watts per square centimeter, such as between about
10.sup.5 watts per square centimeter and about 10.sup.8 watts per
square centimeter, or such as on the order of 10.sup.7 watts per
square centimeter.
[0051] FIG. 3 is a schematic diagram illustrating a proposed
mechanism of etching of SiN.sub.x, assisted by chloride ions, in
accordance with an embodiment of the invention. Without wishing to
be bound by any specific theory, it is proposed that, when Si-rich
regions of SiN.sub.x absorb the laser irradiation, the
electron-poor photoactivated Si can attract chloride ion (Cl.sup.-)
binding, thereby promoting Si--Si bond breaks, which lead to
eventual oxidation. The resulting amorphous oxide is soluble at
high temperatures, leading to thinning of the SiN.sub.x membrane.
It will be appreciated that an analogous mechanism may be present
when other light absorbing materials and electron donor species are
used.
[0052] In accordance with embodiments of the invention, the "light
absorbing material" can include any material that absorbs light in
any of a wide variety of regions of the electromagnetic spectrum,
including ultraviolet wavelengths, such as between about 10 nm and
about 400 nm, and visible wavelengths, such as between about 400 nm
and about 700 nm. The light absorbing material can, for example,
comprise silicon; and can comprise at least one of a
silicon-containing nitride and a silicon-containing carbide. The
light absorbing material can, for example, comprise a chemical
formula SiN.sub.x, where x is greater than or equal to 0 and less
than or equal to 2.
[0053] In accordance with embodiments of the invention, the
"electron donor species" is a chemical entity that donates
electrons to another compound. For example, the electron donor
species can be at least one of a halogen ion and a hydroxide ion.
In one example, the electron donor species can comprise a chloride
ion.
[0054] In accordance with embodiments of the invention, the solvent
can be any suitable solvent that includes the electron donor
species that is used in the method of etching. For example, a
solution of potassium chloride (KCl) can be used to deliver
chloride ions as the electron donor species. It will be appreciated
that a wide variety of solvents, including halogen-containing
solvents such as sodium fluoride, sodium chloride, sodium bromide,
sodium iodide, potassium fluoride, potassium chloride, potassium
bromide, potassium iodide, can be used. Basic solvents can be used
to deliver hydroxide ions as the electron donor species, such as,
for example, potassium hydroxide (KOH), sodium hydroxide (NaOH),
and other basic solvents. In some embodiments, pure water can be
used to deliver the hydroxide ions. Other solvents containing
electron donor species can be used, such as dimethyl sulfoxide
(DMSO) and dimethylformamide (DMF). The solvent can be
biocompatible, and the etching can be performed in a biocompatible
environment. As used herein, "biocompatible" is the quality of not
having toxic or injurious effects on biological systems.
[0055] FIG. 4 is a schematic diagram illustrating a method for
three-dimensional etching, in accordance with an embodiment of the
invention. In FIG. 4, parallel surface photoetching is performed
using a broad light source that reflects from a digital micromirror
device (DMD) 414, or any other spatial light modulator (SLM). This
method can be used for three-dimensional micro-nanofabrication.
More generally, methods in accordance with an embodiment of the
invention can include exposing the interface between the solvent
and the light absorbing material to the light, in the absence of a
lithographic mask. For example, in FIG. 4, no lithographic mask is
between the digital micromirror device 414 and the light absorbing
material 402, which in this example is silicon nitride (SiN). In
the embodiment of FIG. 4, the method includes performing a
three-dimensional, layer by layer etching of the light absorbing
material. A light image projection device (not shown) is used to
provide the light 408, to perform the three-dimensional, layer by
layer etching 418, based on an electrical signal 416 comprising a
three-dimensional etching pattern having features 412a with
dimensions less than 1 micron in size. The beam of light 408 can be
reflected from the spatial light modulator, such as the digital
micromirror device 414, onto the light absorbing material 402. This
technique can be used to perform single step micro-nano
fabrication. Fabrication of an elaborate micro-nanostructure on a
surface, such as a SiN.sub.x surface, would require a
time-consuming serial scanning of the beam. However, using laser
image projection such as a digital micro-mirror device 414, the
design of the etching pattern 416 allows one to fabricate a 3D
scaffold in a layer-by-layer manner at a wafer scale. A location of
the etched feature 412b on the light absorbing material can be
directly controlled, based on the location of the light 408 on the
light absorbing material 402.
[0056] In another embodiment, the etched feature can be formed
underneath a surface of a structure of which the light absorbing
material forms at least a portion. For example, the light 408 can
be transmitted through a surface of a structure (in the position of
coverslip 452 of FIG. 4, for example, but attached to light
absorbing material 402), of which the light absorbing material 402
forms a part. The etching can then proceed underneath the surface
of the structure, on the light absorbing material 402.
[0057] FIG. 5 is a schematic diagram illustrating use of a method
of etching in accordance with an embodiment of the invention to
polish a surface of a light absorbing material, such as to polish a
rough surface of a silicon material. Since light illumination on a
rough surface creates a strong localized light spot 520 on a
sharp-edged surface, this condition accelerates the etching speed
on the sharp-edged surfaces as compared with the surrounding smooth
surfaces, leading to a polished surface 522.
[0058] In accordance with an embodiment of the invention, the
method of etching can include forming a nanopore through the light
absorbing material. For example, the nanopore can have a diameter
greater than about 1 nanometer and less than about 1 micron; and
the light absorbing material can have a thickness less than about 1
micron at the location of the nanopore. In addition, the method can
include using the etching to form an array of a plurality of
nanopores in the light absorbing material. FIG. 6 is a schematic
diagram illustrating components that can be used to control
formation of a nanopore, in accordance with an embodiment of the
invention. In this example, a light source 624, such as a
collimated 532 nm laser, is focused on a SiN.sub.x membrane, or
other light absorbing material 602, using an objective lens, such
as a 60.times. inverted objective lens, 626. The SiN.sub.x membrane
602 is mounted between two fluidic chambers (here, the top referred
to as -cis, the bottom as -trans). (Here, it is noted that the
solvent containing the electron donor species, which is in the
fluid chambers on either side of the light absorbing material 602,
is not shown in FIG. 6). The laser power is electrically controlled
using an electro-optical modulator 628. The current across the
nanopore 630 is monitored by applying a bias voltage 632 between
Ag/AgCl electrodes (the voltage is applied to the -cis chamber in
this scheme). Forming the nanopore can include limiting a power
density of the light after a current measured through the light
absorbing material rapidly increases above a current threshold
during the etching. For this purpose, a threshold detection circuit
634 can be used, so that when a measured current across the
nanopore 630 exceeds the threshold current setpoint of the
threshold detection circuit 634, a trigger circuit 636 provides an
electrical signal to limit or shut off the power of the laser, for
example by controlling the electro optical modulator 628. The
method can include limiting the current threshold to determine the
size of the nanopore 630 formed by the etching; for example, a
larger threshold current can be used to create a larger nanopore,
and a smaller threshold current can be used to create a smaller
nanopore. In addition, the method can include controlling a
thickness of the etched feature using feedback from an optical
measurement of the etched feature. In one example, reflected light
from the light source 624, transmitted lamp light, and fluorescence
from the nanopore 630 (or other etched feature) can be passed
through a dichroic mirror 638, and be detected by a light detection
device 640 such as a CCD camera or spectrometer. Using feedback of
the detected fluorescence, in one example, the thickness of the
etched feature can be controlled by limiting or shutting off the
etching when the desired thickness is reached.
[0059] FIG. 7 is a graph illustrating measurements during a method
of limiting the power density of the light after a current measured
through the light absorbing material rapidly increases above a
current threshold during the etching, in accordance with an
embodiment of the invention. For example, an apparatus such as that
of FIG. 6 can be used to implement such a method. In FIG. 7, the
measured current 742 rapidly increases over a short time frame 744
(for example, less than about 0.5 seconds), to a level above the
threshold current 746. This rapid increase is taken as signifying
the formation of the nanopore (or other feature) in the light
absorbing material. After the current increases above the current
threshold in this fashion during the etching, the power density of
the light is then limited, as discussed above in connection with
FIG. 6.
[0060] In another embodiment, the method can include assisting the
etching by applying a voltage across the light absorbing material
to promote dielectric breakdown of the light absorbing material.
For example, a bias voltage 632 (see FIG. 6) can be set a
sufficiently high level (for example, higher than 100 mV, 200 mV or
300 mV), and be used while applying the light source to the light
absorbing material, in the presence of the solvent containing the
electron donor species, to promote dielectric breakdown of the
light absorbing material in order to assist the etching.
[0061] In another embodiment, the etched feature can define a
nanopore having a greater diameter on one side of the light
absorbing material than on an opposite side of the light absorbing
material. FIG. 8 is a schematic diagram of a nanopore having a
greater diameter on a -cis side 848 of the nanopore, as compared
with a -trans side 850, formed in accordance with an embodiment of
the invention. The size of the nanopore can be demonstrated using
translocation of DNA through the nanopore from the -cis to the
-trans side and vice versa, as described further in experiments,
herein.
[0062] Further embodiments of the invention are apparatuses that
include a nanopore formed by any of the techniques taught herein,
including an array of such nanopores.
[0063] Experimental: Overview
[0064] Experiments in accordance with an embodiment of the
invention have found that SiN.sub.x can be etched under exposure to
light of .about.10.sup.7 W/cm.sup.2 average intensity, with etch
rates that are influenced by the supporting electrolyte. Combining
this controlled etching with dielectric breakdown, an
electrokinetic process for making pores, nanopores of arbitrary
dimensions as small as 1-2 nm in diameter and thickness can easily
be fabricated. Evidence gathered from biomolecule-pore interactions
suggests that the pore geometries obtained using this method are
more funnel-like, rather than hourglass-shaped. Refined control
over pore dimensions can expand the range of applications of
solid-state nanopores, for example, biopolymer sequencing and
detection of specific biomarkers.
[0065] Experiments in accordance with an embodiment of the
invention have investigated the factors that lead to light induced
etching of SiN.sub.x, focusing on photoexcitation, heating, and
chemical catalysis as the driving factors. High temperatures have
been shown to be obtained at the pore under laser irradiation,
depending on the surrounding environment and the laser power. It
has also been shown that coating of the SiN.sub.x with a conformal
layer of HfO.sub.2 effectively eliminates etching under
illumination, suggesting that the solid-electrolyte interface is
required for optically induced SiN.sub.x etching. Next, it has been
demonstrated that SiN.sub.x etching rates are affected by the type
of salt present in the contacting electrolyte, with chloride being
the most effective neutral-pH etch catalyst and hydroxide being a
very fast etchant. It has been found that controlled etching under
applied bias and laser/voltage feedback drives the dielectric
breakdown of nanopores with small diameters (<5 nm) and tunable
effective thickness (0.5-8 nm). Supported by observations of
asymmetric molecular transport across these nanopores, it is
conjectured that the pores are asymmetric in shape, exhibiting a
funnel-like geometry. This approach can serve as a foundation for
high-resolution pores that are comparable in geometry to
protein-based nanopores used in DNA sequencing.
[0066] Experiment: Controlled Photothermal Etching of SiN.sub.x
[0067] FIGS. 9A-9C are diagrams illustrating controlled
photothermal etching, in accordance with an embodiment of the
invention.
[0068] FIG. 9A is a continuous current trace of a SiN.sub.x and
HfO.sub.2-coated SiN.sub.x pore at T.sub.pore=60.degree. C. for 10
min (V=100 mV) under 532 nm laser irradiation (laser turned on
after 5 s), in an experiment in accordance with an embodiment of
the invention. Line 954: A 6.4 nm diameter SiN.sub.x pore with 7 nm
effective thickness. Line 956: A 6.2 nm diameter of
HfO.sub.2-coated SiN.sub.x pore with 5.3 nm effective thickness.
Inset top: I-V curves of HfO.sub.2-coated SiN.sub.x and SiN.sub.x
pore before and after the 10 min laser illumination. For the
HfO.sub.2-coated SiN.sub.x pore, before and after conductance
values were 17 nS and 20 nS, respectively. For the SiN.sub.x pore,
before and after conductance values were 14 nS and 32 nS,
respectively. Expanded current traces at 0-9 s, showing laser
turn-on times, are shown in the inset.
[0069] FIG. 9B is a TEM image of 75 nm-thick SiN.sub.x membrane
after 47 mW laser illumination for 10 min in water, 0.4 M sodium
acetate, and 0.4 M KCl (T=300.degree. C.), in an experiment in
accordance with an embodiment of the invention. Membrane
thicknesses at each point are related to the image brightness, and
intensity values for the different buffers were 181, 37, and 49 for
KCl, sodium acetate, and water, respectively.
[0070] FIG. 9C is a TEM image of a 75 nm-thick SiN.sub.x membrane
after 1.5 mW laser illumination for 8 h in 4 M KCl, in an
experiment in accordance with an embodiment of the invention.
[0071] Previous experiments in accordance with an embodiment of the
invention found that high temperatures are reached when SiN.sub.x
is irradiated using a focused 532 nm laser at high power densities
(.about.3.times.10.sup.7 W/cm.sup.2 at 50 mW laser power, assuming
a 1/e.sup.2 beam diameter of 710 nm). To investigate the impact of
the high-energy beam incident upon SiN.sub.x on its integrity, in
FIG. 9A there is shown the ion current through a 6.4 nm diameter
SiN.sub.x nanopore fabricated using a TEM prior to the experiment
(trace 954), when at t=5 s, a 30 mW laser beam has been switched
on. The instantaneous jump in ion current (see inset) is due to
rapid heating caused by the laser, which results in a peak
temperature at the pore of T.sub.p=60.degree. C. Pore temperature
was calculated using the ion current enhancement profile, as
described in our previous report (5). However, following this
current jump, we observed a gradual increasing of the ion current,
and over a period of 10 min, the pore current approximately
doubled. This ion current enhancement is likely due to a
combination of SiN.sub.x thinning and pore expansion, since both
processes are consistent with stimulated dissolution of the
SiN.sub.x, as recently observed using a similar-wavelength (488 nm)
and similar-intensity laser. (2, 5, 6). When a bare 7.6 nm diameter
SiN.sub.x pore surface was passivated with a conformal layer of 1
nm-thick HfO.sub.2, a dielectric material, it was found that
HfO.sub.2 greatly inhibits pore dissolution (trace 956). The
current changes for the bare and HfO.sub.2-coated SiN.sub.x pores
are 5.0 and 0.6 pA/s, respectively. I-V curves of both pores before
and after the 10 min laser treatments, shown in the inset,
highlight the photoreactivity of bare SiN.sub.x.
[0072] To gain more insight into the factors that influence the
observed SiN.sub.x photothermal reactivity, FIG. 9B shows
bright-field TEM images of a membrane which has been irradiated for
10 min at different locations using a 47 mW power
(T.sub.p=300.degree. C.), where prior to irradiation, the cis and
trans fluids were replaced to expose the SiN.sub.x membrane to
either 0.4 M KCl, 0.4 M sodium acetate or pure water. Brightness in
this TEM image corresponds to a higher transmitted electron dose
through the membrane, which means that the SiN.sub.x is thinnest in
the case of chloride exposure. Based on these measurements,
chloride is a more effective promoter of SiN.sub.x dissolution,
whereas in acetate and pure water, SiN.sub.x dissolution is
minimal. This series of experiments was repeated with very similar
results. Further, FIG. 9C shows the impact of 8 h laser
illumination at a power of 1.5 mW in 4 M KCl buffer. The slow
etching of SiN.sub.x under a weak laser illumination, in which the
photothermal effect is negligible, emphasizes that the etching
process is photothermally activated. (3, 4).
[0073] Experiment: SiN.sub.x Dissolution Kinetics and Mechanism
[0074] FIG. 10A is a schematic diagram of photochemical etching on
freestanding SiN.sub.x membrane, in an experiment in accordance
with an embodiment of the invention. When a collimated 532 nm laser
was focused on a SiN.sub.x freestanding membrane, locally
photochemical etching starts at both sides of the membrane due to
its exposure in aqueous chloride solutions. Optical image after
photochemical etching with P=47 mW (diameter of light spot: 710 nm)
and V=1 V for 3 min in 0.4 M KCl (etch width fwhm: 533 nm, maximum
etch depth: 26.2 nm).
[0075] FIG. 10B shows AFM-measured etch depth profiles, laser at
different laser powers and times, for 75 nm-thick SiNx supported by
2.5-.mu.m-thick silicon oxide on silicon, in an experiment in
accordance with an embodiment of the invention. Inset: AFM image of
an etched array at indicated laser powers and times. The distance
between each etched point was 3 .mu.m. White lines are depth
profiles measured across the center of the etch regions.
[0076] FIG. 3 (above) shows a proposed mechanism of photochemical
SiN.sub.x etching assisted by chloride ions, in accordance with an
embodiment of the invention. When Si-rich regions of SiN.sub.x
absorb the laser irradiation, the electron-poor photoactivated Si
can attract Cl.sup.- binding, promoting Si--Si bond breaks which
lead to eventual oxidation. The resulting amorphous oxide is
soluble at high temperatures, leading to thinning of the SiN.sub.x
membrane.
[0077] FIG. 11 is a continuous current trace of 75 nm-thick
SiN.sub.x membrane etching in 0.4 M KCl and 4 M KCl using a 10 mV
probing voltage (laser power was 47 mW), in an experiment in
accordance with an embodiment of the invention. When current
threshold levels of 1 nA and 12 nA were reached for 0.4 and 4 M
KCl, respectively, laser irradiation was shut off. Inset left:
Optical images before laser-assisted SiN.sub.x thinning, after
first pore at 0.4 M KCl, and after second pore at 4 M KCl,
respectively (left-to-right). Inset right: I-V curves after first
and second pore formation, both recorded in 4 M KCl.
[0078] In more detail, experiments in accordance with an embodiment
of the invention studied the dependence of light power and
electrolyte strength on SiN.sub.x dissolution kinetics by
quantitative measurements using atomic force microscopy (AFM). FIG.
10A shows the impact of a 3 min irradiation on a freestanding 75
nm-thick SiN.sub.x membrane. AFM image of 47 mW laser illumination
with V=1 V for 3 min yields Gaussian distributed etch profiles (1)
(etch width fwhm: 533 nm, maximum etch depth: 26.2 nm). Notably,
this etch profile matches the laser beam profile, as previously
probed using nanopore tomography. (7). Further, it was found that
the etch rate depends nonlinearly on the laser power, which points
to an activated chemical process as suggested by an Arrhenius
equation, as shown in FIG. 10B. The combined observations suggest
the following mechanism for the observed etching process: when
silicon rich SiN.sub.x (x.about.1) absorbs blue-green light,
silicon bonds are activated by charge separation, which can likely
lead to destabilization of Si--Si bonds in the Si-rich material.
(4). It is hypothesized that Cl.sup.- can stabilize the transiently
formed activated Si by forming transient Si--Cl bonds, which can
explain Cl.sup.--promoted breakdown of activated Si--Si bonds, as
shown in FIG. 3. Noted here is a known process in which chlorine
plasma efficiently breaks down Si.sub.3N.sub.4: (8)
Cl+Si.sub.3N.sub.4.fwdarw.SiCl.sub.4+NCl.sub.3, N, N.sub.2 (1)
[0079] Interestingly, photothermal heating has been shown to
accelerate this process, especially at 300-500.degree. C. (9) While
the formed Si--Cl bond is unstable in the presence of water,
Cl.sup.- can be rapidly displaced by water molecules leading to
oxidation, in accordance with the observed reduction in PL upon
irradiation. (2) Dissolution of the resulting amorphous SiO.sub.2
product can then be explained by the hot spot induced by the laser,
since solubility of amorphous SiO.sub.2 is enhanced greatly by
temperature. (10) Further evidence of chloride mediated SiN.sub.x
etching was obtained when 75 nm SiNx etching time was compared in
0.4 and 4 M KCl on the same membrane. FIG. 11 shows a current trace
of a membrane under a focused 47 mW laser illumination at a low
probing voltage of V=10 mV, in which a hole was formed after 170 s
when the electrolyte was 0.4 M KCl. In contrast, using 4 M KCl, a
hole formed after 42 s, suggesting that an increased Cl.sup.-
concentration is accelerating pore formation. Similar experiments
using 0.04 M KCl resulted in .about.3 times slower hole formation,
consistent across three independent experimental sets. Finally,
while small pH decreases (1-2 units) do result from heating tris
buffer, (11) it is noted that etching was even faster at a higher
pH, suggesting that slightly lower pH values cannot be responsible
for this etching mechanism (0.1 M KOH).
[0080] Experiment: Laser-Assisted Dielectric Breakdown of
Nanopores
[0081] FIGS. 12A and 12B are diagrams illustrating laser-assisted
dielectric breakdown for nanopore fabrication, in experiments in
accordance with an embodiment of the invention.
[0082] FIG. 12A shows a continuous current trace obtained during
SiN.sub.x laser assisted dielectric breakdown (35 mW laser power,
V=1 V, laser turned on at 0 s), in an experiment in accordance with
an embodiment of the invention. When pore formation is indicated by
the current reaching 0.5 nA, the software turned off the laser, and
a probing voltage of V=50 mV was applied. Inset (left): I-V curve
of 2.4 nm diameter pore with 3 nm effective thickness formed using
this method. Inset (right): 9 min continuous trace during pore
expansion and stabilization at 50 mV probing voltage.
[0083] FIG. 12B is a scatter plot of pore thicknesses vs breakdown
times for 29 pores with three different laser powers used (P=47 mW
shown in red, P=35 mW shown in blue, and P=24 mW shown in green),
in an experiment in accordance with an embodiment of the invention.
Squares represent one standard deviation the parameters. Average
breakdown times were 165.+-.67 s (47 mW), 536.+-.288 s (35 mW), and
814.+-.440 s (24 mW), and average effective respective thicknesses
were 1.8.+-.1.1 nm, 3.9.+-.1.1 nm, and 4.6.+-.2.3 nm.
[0084] In more detail, it is found that laser-assisted SiN.sub.x
etching greatly facilitates the fabrication of ultrathin and
ultrasmall nanopores using dielectric breakdown. FIG. 12A shows a
continuous current trace during laser-assisted dielectric breakdown
(V=1 V applied to the cis chamber, P=47 mW). When the SiN.sub.x
membrane is sufficiently thin, breakdown can occur (<8 nm), and
pore formation is marked by a sharp current rise, as seen at 180 s
in the trace. Using software-based feedback, we stop the high-power
laser illumination and reduce the voltage to 50 mV when the ion
current reaches our specified threshold current (0.5 nA). After
initial pore formation, we apply a weak voltage (50-100 mV) for up
to 30 min to allow equilibration and stabilization of the pore
current prior to adding biomolecules (see FIG. 12A, inset). Since
both laser heating and voltage were used, it is assumed that the
dielectric breakdown process includes SiN.sub.x thinning, thermal
heating acceleration of defect production, and additional electric
field by light. (1, 12-14) Pore size and its effective thickness
are calculated using established methods that measure DNA blockades
and DNA diameters (2.2 nm for dsDNA and 1.2 nm for ssDNA). (15,
16). Laser induced breakdown allows fabrication of a sub-5 nm
diameter pore with thickness values that are well below 5 nm, as
indicated by high blockade amplitudes observed for translocations.
In FIG. 12B are shown pore thicknesses vs breakdown times and
resulting pore diameters obtained using three different laser
powers for a total of 29 pores made. The boxes represent one
standard deviation in each parameter for the experiments conducted.
It is found that weak laser powers produce larger deviations in
breakdown times and thicker pores, as compared with high laser
powers, for which pores are routinely obtained with .about.2 nm
average thickness within 1-2 min of processing. The results
demonstrate the sophisticated control that laser power and voltage
have over the delicate size and shape of nanopores.
[0085] Experiment: Evidence for Asymmetric Nanopore Shape in
Laser-Assisted Breakdown Pores
[0086] FIG. 12C is a diagram showing typical .DELTA.I/I0 vs log
t.sub.d scatter plots obtained for 2.5 kbp DNA translocation
dynamics through a 3.3 nm diameter pore with 2.8 nm effective
thickness at 300 mV from cis (left) and trans (right) side, in an
experiment in accordance with an embodiment of the invention.
Right: concatenated traces of DNA translocation events,
low-pass-filtered at 10 kHz. DNA concentrations at cis and trans
sides were 6 nM and 4 nM, respectively.
[0087] One aspect of these pores fabricated using high laser powers
is their shape, which is extremely difficult to image, yet for
which DNA translocation data suggests a consistent pattern. In FIG.
12C, there is shown 2.5 kbp DNA translocation dynamics through a
pore (d=3.3 nm, t.sub.eff=2.8 nm) made by the method, as a function
of the translocation direction. In the cis-to-trans direction, DNA
translocation events have to clear two-blockade levels, a shallow
one and a deep one, which corresponds to docking at the pore
entrance due to its funnel-like shape, and translocation.
Interestingly, this docking time is very long and almost always
ends with translocation. In contrast, in the trans-to-cis direction
one observes rapid deep events, suggesting that DNA docking times
are much faster in this direction. A hypothesized mechanism for
this result, which is observed consistently for pores generated
using this method, is that the pore shape is asymmetric and
funnel-like in shape. As the cartoons illustrate (see insets to
FIG. 12C, right), this asymmetric pore structure yields a pocket
for DNA that results in longer docking times than when DNA
approaches from the narrower opening. Further, different capture
rates are found for DNA entering from either side, with rates of
3.19.+-.0.08 s.sup.-1 nM.sup.-1 for cis-to-trans direction and
1.05.+-.0.03 s.sup.-1 nM.sup.-1 for trans-to-cis direction. Both of
these features, namely longer docking times and faster capture
rates from cis entry into pore than from trans entry, were
previously observed for ssDNA entry into the protein channel
a-hemolysin. (17, 18) The asymmetric geometry is also suggested
from the corresponding increase of effective thickness as pore
size.
[0088] Ultrathin nanopores fabricated using laser-assisted
dielectric breakdown can be used to differentiate two dsDNA lengths
based on their dwell times in a DNA mixture. (19, 20) FIGS. 13A and
13B show discrimination of 250 bp and 2.5 kbp DNA, in experiments
in accordance with an embodiment of the invention. FIG. 13A shows
continuous 1 s trace excerpts at 200, 250, and 300 mV for a 2.7 nm
pore with 1.7 nm effective thickness for a mixture of 250 bp (30
nM) and 2.5 kbp (3 nM) dsDNA added to the cis chamber. Traces were
low-pass filtered at 20 kHz. FIG. 13B shows .DELTA.I/I0 vs log
t.sub.d of a mixture of 250 bp and 2.5 kbp at 200, 250, and 300 mV,
showing an optimal resolution between the two lengths at 300
mV.
[0089] In more detail, FIG. 13A shows continuous current traces for
a mixture of 250 bp (30 nM) and 2.5 kbp (3 nM) translocating
through an ultrathin pore (d=2.7 nm, t.sub.eff=1.7 nm) at 200, 250,
and 300 mV. Scatter plots of .DELTA.I/I0 vs log t.sub.d, shown in
FIG. 13B, reveal mean log t.sub.d values for 250 bp/2.5 kbp of
3.80.+-.0.01/4.79.+-.0.01, 3.36.+-.0.03/4.47.+-.0.02, and
3.19.+-.0.05/4.29.+-.0.02 .mu.s for 200, 250, and 300 mV,
respectively. In this experiment, expansion of the pore was minimal
despite the ultrathin SiN.sub.x membrane. Also, it is found that as
voltage increases, the 2.5 kbp dsDNA produces larger fractional
blockades (but not the 250 bp DNA), attributed to interactions of
the longer DNA coil with significant electric fields outside the
pore. (20) It is noted that the long docking times observed with
the larger 3.3 nm diameter pore in FIG. 12 was not observed in this
experiment, because the pore diameter is significantly smaller,
which yields an electric field outside the pore that is too weak to
trap DNA in the pore mouth.
[0090] Experiment: DNA Homopolymer Differentiation Using Sub-2
nm-Thick Pores
[0091] FIGS. 14A-14D are diagrams showing current discrimination of
dC and dA homopolymers using an ultrathin laser-assisted breakdown
pore, in experiments in accordance with an embodiment of the
invention.
[0092] FIG. 14A is a schematic diagram of sequentially unzipping
dsDNA to expose a predetermined temporal sequence of homopolymers
to the pore, in an experiment in accordance with an embodiment of
the invention. During unzipping DNA duplex, either a poly(dC) or a
poly(dA) portion of the ssDNA template resides inside the pore,
allowing residual ion currents to be probed. Designed DNA is in the
probing order of poly(dC), poly(dA), and poly(dC).
[0093] FIG. 14B is a current trace of DNA unzipping in a 1.4 nm
diameter pore with 1.8 nm effective thickness at 200 mV (trace was
low-pass-filtered at 1 kHz), in an experiment in accordance with an
embodiment of the invention. The expanded trace with poly(dC) and
poly(dA) labels is shown in the inset.
[0094] FIG. 14C shows I levels of poly(dC) (blue markers) and
poly(dA) (red markers) and .DELTA.I dA-dC (black squares), in an
experiment in accordance with an embodiment of the invention.
Current levels were obtained by double Gaussian fits to the current
histograms of each event.
[0095] FIG. 14D is a histogram of differential .DELTA.I dA-dC with
the Gaussian fit, in an experiment in accordance with an embodiment
of the invention. Mean .DELTA.I current difference between dA and
dC levels was 16.9.+-.1.4 pA.
[0096] In more detail, in FIGS. 14A-14D, there are shown the
results of experiments involving DNA homopolymer differentiation
using sub-2 nm thick pores. These experiments demonstrate the
ability of an ultrathin sub-2 nm diameter pore (d=1.4 nm,
t.sub.eff=1.8 nm) to discriminate among two homopolymer types. FIG.
14A schematically illustrates a vertically oriented membrane where
a long template DNA (64-nucleotide) is inserted into the pore, and
hybridized DNA oligomers (15 and 19 nucleotides) are sequentially
unzipped by voltage-induced force applied to the DNA template. In
this experiment, the current levels of poly(dA) and poly(dC) that
are in the pore during the three-stage unzipping process are
probed. Under an applied voltage (200 mV), the poly(dC) tail of the
template DNA enters the pore and resides there until the first
duplex unzips, then the poly(dA) resides in the pore, and finally
the poly(dC) resides in the pore. It was found that events longer
than 5 ms comprised 3.36% of the total events for the
single-stranded 64-nucleotide DNA template, whereas for the
hybridized sample, 20.3% of the events lasted longer than 5 ms,
which is attributed to unzipping events. (21) Within this long
event population, in 18.6% of the events, there were observed two
distinct blockade levels, which are ascribed to the helix structure
of poly(dC) and poly(dA) (FIG. 14B). As with .alpha.-hemolysin
experiments, the 1.3 nm-wide poly(C) helical secondary structure
can be accommodated in 1.4 nm pore, while the 2.1 nm-wide poly(A)
helix is too large to fit inside the narrow pore, (22) resulting in
a shallower blockade current for poly(dA).
[0097] FIG. 14C shows current blockade levels of poly(dC) and
poly(dA) and its difference from the same pore. The mean .DELTA.I
difference between dA and dC (FIG. 14D) in all these events is
16.9.+-.1.4 pA, in good agreement with previous .alpha.-hemolysin
and MspA experiments. (22-24) Although nucleotide resolution using
a solid-state nanopore was previously demonstrated by homopolymer
discrimination, (25-27) these results show that ultrathin
solid-state nanopores have the sensitivity to discriminate both
types of polymers within a single molecular chain.
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[0125] The teachings of all patents, published applications and
references cited herein are incorporated by reference in their
entirety.
[0126] While example embodiments have been particularly shown and
described, it will be understood by those skilled in the art that
various changes in form and details may be made therein without
departing from the scope of the embodiments encompassed by the
appended claims.
Sequence CWU 1
1
3115DNAArtificial Sequencehybridized DNA oligomers 1tttttttttt
ggggg 15219DNAArtificial Sequencehybridized DNA oligomers
2cccttgatgc actggctta 19364DNAArtificial Sequencelong template DNA
3aaaaaaaaaa cccccccccc gggaactacg tgaccgaata aaaaaaaaac cccccccccc
60cccc 64
* * * * *