The subject matter described herein relates to methods, devices, and systems for sequencing nucleic acid polymers.
Nucleic acid bases can be read by using electron tunneling current signals generated as the nucleotides pass through a tunnel gap functionalized with adaptor molecules. For example, PCT publication nos. WO2009/117522A2, WO 2010/042514A1, WO 2009/117517, and WO2008/124706A2, U.S. publication nos. US2010/0084276A1, and US2012/0288948, are all hereby incorporated by reference herein in their entireties. Conventionally, bases have been read using gold electrodes functionalized with adaptor molecules. Carbon nanotubes functionalized with adaptor molecules have also been described for use as electrodes in PCT publication nos. WO2009/117517 and WO 2010/042514A1, and U.S. publication nos. US2011/0168562 and US2011/0120868, which are incorporated herein by reference in their entireties.
While gold has been found to work well as an electrode material, it suffers from limitations. For examples, it is often incompatible with current technologies used for fabricating electronic devices, owing to its rapid diffusion in silicon and its propensity to form deep level traps, reducing minority carrier lifetime. Second, the tunneling signals generated by the most successful adaptor molecule tried to date, i.e., (4(5)-(2-mercaptoethyl)-1H imidazole-2-carboxamide), can have a large background generated by water alone. This is illustrated in
In view of the foregoing, it would be desirable to provide improved methods, devices, and systems for sequencing nucleic acid polymers. In one aspect according to some embodiments, methods, devices, and systems for sequencing nucleic acid polymers are provided that utilize an electrode material, functionalized with one or more adaptor molecules, that is compatible with semiconductor fabrication processes. In another aspect according to some embodiments, methods, devices, and systems for sequencing nucleic acid polymers are provided that utilize an electrode material, functionalized with one or more adaptor molecules, that is capable of generating signals from DNA nucleobases without interference from water signals. One or both of these improvements and advantages, and/or other improvements and advantages, can be provided in accordance with the present disclosure.
Embodiments of the subject matter described herein provide methods, devices, and systems for sequencing nucleic acid polymers.
For example, some embodiments of the present disclosure provide methods, devices, and systems for sequencing nucleic acid polymers that utilize palladium (Pd), at least in part (e.g., whether it be pure palladium, a palladium alloy, or other composition comprising palladium), as an electrode material that is (i) functionalized with one or more adaptor molecules and (ii) capable for use to sense one or more chemical compositions.
In some embodiments, a device for identifying a chemical composition (e.g., single molecules) and a corresponding method of fabricating the device are provided. The device includes a first electrode and a second electrode separated from the first electrode by a dielectric material (e.g., dielectric material having about 1 to 5 nm thickness). The first electrode, second electrode, or both have at least one adaptor molecule chemically tethered thereto. In some embodiments, at least one of the first electrode and the second electrode comprises palladium metal (e.g., pure palladium or a palladium alloy). In some embodiments, the adaptor molecule comprises 4(5)-(2-mercaptoethyl)-1H imidazole-2-carboxamide. In some embodiments, the adaptor molecule comprises 4H-1,2,4-triazole-3-carboxamide. In other embodiments, the adaptor molecule comprises 2-(2-carbamoyl-1H-imidazol-4-yl)ethylcarbamodithioate.
In an embodiment, an apparatus and corresponding method for sensing a chemical composition are provided. For example, in some embodiments, a nucleic acid base is caused to pass through a tunnel gap having electrically-separated electrodes, where at least one of the electrically-separated electrodes comprises palladium metal functionalized with an adaptor molecule. A type of the nucleic acid base is identified based on a tunneling current generated as a result of the nucleic acid base passing through the tunnel gap. In some embodiments, the adaptor molecule comprises 4(5)-(2-mercaptoethyl)-1H imidazole-2-carboxamide. In some embodiments, the adaptor molecule comprises 4H-1,2,4-triazole-3-carboxamide. In other embodiments, the adaptor molecule comprises 2-(2-carbamoyl-1H-imidazol-4-yl)ethylcarbamodithioate.
In some embodiments, a device for identifying one or more molecules (e.g., single molecules) is provided and comprises a first electrode, a second electrode separated from the first electrode by a dielectric material of about 1 to about 5 nm thickness, at least one adaptor molecule chemically tethered to the first electrode, and at least one adaptor molecule chemically tethered to the second electrode. In some embodiments, at least one of the first electrode and the second electrode comprises palladium metal.
In some embodiments, both of the first electrode and the second electrode comprise palladium metal. In some embodiments, at least one of the first electrode and the second electrode comprise an alloy of palladium. In some embodiments, at least one adaptor molecule tethered to the first electrode, the at least one adaptor molecule tethered to the second electrode, or both comprise 4(5)-(2-mercaptoethyl)-1H imidazole-2-carboxamide.
In some embodiments, at least one adaptor molecule tethered to the first electrode, the at least one adaptor molecule tethered to the second electrode, or both comprise 4H-1,2,4-triazole-3-carboxamide.
In some embodiments, the at least one adaptor molecule tethered to the first electrode, the at least one adaptor molecule tethered to the second electrode, or both comprise 2-(2-carbamoyl-1H-imidazol-4-yl)ethylcarbamodithioate.
In some embodiments, the electrodes are held under potential control with respect to reference electrode. In some embodiments, the potential of the palladium surface is maintained at between about +0.5V and about −0.5V vs. Ag/AgCl.
In some embodiments, an apparatus for sensing a chemical composition is provided and may comprise means for causing a nucleic acid base to pass through a tunnel gap having electrically-separated electrodes, where at least one of the electrically-separated electrodes comprises palladium metal functionalized with an adaptor molecule. Such embodiments may also include means for identifying a type of the nucleic acid base based on a tunneling current generated as a result of the nucleic acid base passing through the tunnel gap. Such means may be a computer processor analyzing signal data to determine the identity of the nucleic acid. Such means may also include databases for storing signature signal data for a plurality of molecules to be identified.
In some embodiments, both of the electrically-separated electrodes comprise palladium metal.
In some embodiments, at least one of the electrically-separated electrodes comprises an alloy of palladium.
In some embodiments, the adaptor molecule comprises 4(5)-(2-mercaptoethyl)-1H imidazole-2-carboxamide. In some embodiments, the adaptor molecule comprises 4H-1,2,4-triazole-3-carboxamide, or 2-(2-carbamoyl-1H-imidazol-4-yl)ethylcarbamodithioate.
In some embodiments, a method of fabricating a device capable of sensing a chemical composition is provided and may comprise one or more of the following steps (and in some embodiments, a plurality, and in some embodiments, all steps): providing a first electrode, providing a second electrode separated from the first electrode by a dielectric material of about 1 to about 5 nm thickness, chemically tethering at least one adaptor molecule to the first electrode, and chemically tethering at least one adaptor molecule to the second electrode. In some embodiments, at least one of the first electrode and the second electrode comprises palladium metal.
In some embodiments, such methods may also include at least one of chemically tethering at least one adaptor molecule to the first electrode, chemically tethering at least one adaptor molecule to the second electrode, or both, comprises chemically tethering 4(5)-(2-mercaptoethyl)-1H imidazole-2-carboxamide to the first electrode, second electrode, or both.
In some embodiments, such methods may also include at least one of chemically tethering at least one adaptor molecule to the first electrode, chemically tethering at least one adaptor molecule to the second electrode, or both, comprises chemically tethering 4H-1,2,4-triazole-3-carboxamide to the first electrode, second electrode, or both.
In some embodiments, such methods may include at least one of chemically tethering at least one adaptor molecule to the first electrode, chemically tethering at least one adaptor molecule to the second electrode, or both, comprises chemically tethering 2-(2-carbamoyl-1H-imidazol-4-yl)ethylcarbamodithioate to the first electrode, second electrode, or both.
In some embodiments, a method for sensing a chemical composition is provided and may include one or more of the following steps (in some embodiments, a plurality of such steps, and in some embodiments, all of such steps): causing a nucleic acid base to pass through a tunnel gap having electrically-separated electrodes, where at least one of the electrically-separated electrodes comprises palladium, and identifying a type of the nucleic acid base based on the tunneling current generated as a result of the nucleic acid base passing through the tunnel gap. Such identifying may comprise using computers, processors, and the like, to perform steps of analyzing the signal data to eliminate noise and defects, and/or comparing the signal data to signature signal data for a nucleic acid so as to identify the nucleic acid.
Some embodiments include a computer system for sensing a chemical composition, where the system comprising at least one processor, and where the processor includes computer instructions operating thereon for performing any of the methods taught by the present disclosure.
In some embodiments, a computer program for sensing a chemical composition is provided and comprises computer instructions for performing any of the methods taught by the present disclosure.
In some embodiments, a computer readable medium containing a program is provided, where the program includes computer instructions for performing any of the methods taught by the present disclosure.
The accompanying drawings, which are incorporated in and constitute a part of this specification, show certain aspects of the subject matter disclosed herein and, together with the description, help to explain some of the principles associated with the disclosed embodiments. In the drawings:
Still referring to
Another advantage of probes that include palladium (e.g., pure Pd or Pd alloy) lies with their ability to generate reads from DNA bases at a setpoint conductance that is much smaller than was used for gold electrodes with the 4(5)-(2-mercaptoethyl)-1H imidazole-2-carboxamide adaptor molecules. By way of illustration in accordance with some embodiments, and as shown below, reliable signals are obtained with a tunnel gap of 4 pS conductance, well below the 12 pS that had to be used to acquire the data taken with gold electrodes (
These same conditions also produced copious amounts of signal when nucleotides were added to the tunnel junction.
Operation at this low tunnel conductance provides excellent separation of the signals from the bases.
The device configurations described above in connection with
In various embodiments of the present disclosure, any suitable adaptor molecule(s) can be tethered to the first and/or second electrodes of a device as reading molecules for recognition tunneling. In some embodiments, the adaptor molecule is 4(5)-(2-mercaptoethyl)-1H imidazole-2-carboxamide. In some embodiments, the adaptor molecule is 4H-1,2,4-triazole-3-carboxamide. In some embodiments, the adaptor molecule is 2-(2-carbamoyl-1H-imidazol-4-yl)ethylcarbamodithioate.
Synthesis of the 5-substituted-4H-1,2,4-triazole-3-carboxamide molecule just described is described as follows and in connection with
Still referring to
With further reference to
Still referring to
Preparation of the dithiocarbamate derivative of 4(5)-(2-aminoethyl)-1H-imidazole-2-carboxamide described above, for example, for use as a reading molecule for recognition tunneling is described as follows and in connection with
Tunneling measurements were taken using the adaptor molecules described in connection with
In some embodiments of the present disclosure, palladium electrodes may catalyze a number of chemical reactions. For example, and in particular, in some embodiments, cyclic voltammetry shows that phosphate is strongly adsorbed on the electrodes. Such an effect, in some embodiments, becomes more pronounced upon the potential of the palladium exceeding, for example, about +0.5V (adsorption). In addition, in some embodiments, such an effect becomes less pronounced (i.e., more negative) than about −0.5V (desorption) with respect to an Ag/AgCl reference electrode. Thus, in some embodiments, it may be advantageous to retain the palladium electrodes within such a range of potentials with respect to a reference electrode (for example). In some embodiments, the most negative electrode of the pair may be held more positive than about −0.5V vs. Ag/AgCl and the most positive of the pair, in some embodiments, may be held more negative than about +0.5V vs. Ag/AgCl.
Various implementations of the embodiments disclosed above, in particular at least some of the methods/processes disclosed, may be realized in digital electronic circuitry, integrated circuitry, specially designed ASICs (application specific integrated circuits), computer hardware, firmware, software, and/or combinations thereof. These various implementations may include implementation in one or more computer programs that are executable and/or interpretable on a programmable system including at least one programmable processor, which may be special or general purpose, coupled to receive data and instructions from, and to transmit data and instructions to, a storage system, at least one input device, and at least one output device.
Such computer programs (also known as programs, software, software applications or code) include machine instructions for a programmable processor, for example, and may be implemented in a high-level procedural and/or object-oriented programming language, and/or in assembly/machine language. As used herein, the term “machine-readable medium” refers to any computer program product, apparatus and/or device (e.g., magnetic discs, optical disks, memory, Programmable Logic Devices (PLDs)) used to provide machine instructions and/or data to a programmable processor, including a machine-readable medium that receives machine instructions as a machine-readable signal. The term “machine-readable signal” refers to any signal used to provide machine instructions and/or data to a programmable processor.
To provide for interaction with a user, some of the subject matter described herein may be implemented on a computer having a display device (e.g., a CRT (cathode ray tube) or LCD (liquid crystal display) monitor and the like) for displaying information to the user and a keyboard and/or a pointing device (e.g., a mouse or a trackball) by which the user may provide input to the computer. For example, this program can be stored, executed and operated by the dispensing unit, remote control, PC, laptop, smart-phone, media player or personal data assistant (“PDA”). Other kinds of devices may be used to provide for interaction with a user as well; for example, feedback provided to the user may be any form of sensory feedback (e.g., visual feedback, auditory feedback, or tactile feedback); and input from the user may be received in any form, including acoustic, speech, or tactile input.
Certain embodiments of the subject matter described herein may be implemented in a computing system and/or devices that includes a back-end component (e.g., as a data server), or that includes a middleware component (e.g., an application server), or that includes a front-end component (e.g., a client computer having a graphical user interface or a Web browser through which a user may interact with an implementation of the subject matter described herein), or any combination of such back-end, middleware, or front-end components. The components of the system may be interconnected by any form or medium of digital data communication (e.g., a communication network). Examples of communication networks include a local area network (“LAN”), a wide area network (“WAN”), and the Internet.
The computing system according to some such embodiments described above may include clients and servers. A client and server are generally remote from each other and typically interact through a communication network. The relationship of client and server arises by virtue of computer programs running on the respective computers and having a client-server relationship to each other.
For example, as shown in
Similarly,
Any and all references to publications or other documents, including but not limited to, patents, patent applications, articles, webpages, books, etc., presented in the present application, are herein incorporated by reference in their entirety.
Although a few variations have been described in detail above, other modifications are possible. For example, any logic flow depicted in the accompanying figures and described herein does not require the particular order shown, or sequential order, to achieve desirable results. Other implementations may be within the scope of at least some of the following claims.
Example embodiments of the devices, systems and methods have been described herein. As noted elsewhere, these embodiments have been described for illustrative purposes only and are not limiting. Other embodiments are possible and are covered by the disclosure, which will be apparent from the teachings contained herein. Thus, the breadth and scope of the disclosure should not be limited by any of the above-described embodiments but should be defined only in accordance with claims supported by the present disclosure and their equivalents. Moreover, embodiments of the subject disclosure may include methods, systems and devices which may further include any and all elements from any other disclosed methods, systems, and devices, including any and all elements corresponding to methods, systems and devices for sensing chemical composition. In other words, elements from one or another disclosed embodiments may be interchangeable with elements from other disclosed embodiments. In addition, one or more features/elements of disclosed embodiments may be removed and still result in patentable subject matter (and thus, resulting in yet more embodiments of the subject disclosure).
This application claims benefit under 35 USC §119(e) of U.S. provisional patent application no. 61/620,167, filed Apr. 4, 2012, entitled, “Electrodes for Sensing Chemical Composition” the entire disclosure of which is herein incorporated by reference.
Inventions of the present application were made with government support under NIH Grant No. R01 HG006323, awarded by the National Institute of Health. The U.S. Government has certain rights in inventions disclosed herein.
Number | Date | Country | |
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61620167 | Apr 2012 | US |