Expanding the Molecular Alphabet of DNA-Based Data Storage Systems with Neural Network Nanopore Readout Processing

Abstract

DNA is a promising next-generation data storage medium, but challenges remain with synthesis costs and recording latency. Here, we describe a prototype of a DNA data storage system that uses an extended molecular alphabet combining natural and chemically modified nucleotides. Our results show that MspA nanopores can discriminate different combinations and ordered sequences of natural and chemically modified nucleotides in custom-designed oligomers. We further demonstrate single-molecule sequencing of the extended alphabet using a neural network architecture that classifies raw current signals generated by Oxford Nanopore sequencers with an average accuracy exceeding 60% (39× larger than random guessing). Molecular dynamics simulations show that the majority of modified nucleotides lead to only minor perturbations of the DNA double helix. Overall, the extended molecular alphabet may potentially offer a nearly 2-fold increase in storage density and potentially the same order of reduction in the recording latency, thereby enabling new implementations of molecular recorders.

A nanopore interface for higher bandwidth DNA computing

“…13 ). Future work will be aimed at: 1) further expansion of the barcode space, for example, with chemical modifications to the DNA that could expand the dynamic range of barcode signal space 40 , and/or the ability to read sequential barcode regions within a single output strand 25 , 41 , 42 ; and 2) increasing the reaction speed and sensitivity of DSD reactions, for example by spatially-localizing DNA components to the nanopore sensor membrane 43 , 44 . Increasing the scale and speed of our detection strategy for more complex molecular computing architectures, such as cascaded circuits or oscillators, will further take advantage of our method’s ability to generate both multiplexed and kinetic readouts.…”

Emerging Approaches to DNA Data Storage: Challenges and Prospects

“…During the 2010s, extensive innovations in algorithm development have enabled reliable storage of data even under significant errors. Grass et al used modern error-correcting codes in the context of DNA storage, and a variety of different schemes have been proposed. ,− ,, While physical and logical redundancy lower the storage density of DNA, recent works have proposed to raise it by expanding the DNA alphabet using composite natural letters ,, or chemically modified nucleotides …”

“…(Figure A,B). , As an alternative, sequencing using protein nanopores, commercialized by Oxford Nanopore Technologies, has been used because of its ease of implementation, automation, and portability (Figure C). ,, Nanopore sequencing uses electrical readouts rather than fluorescence detection to identify each base of a DNA strand as it moves through a biological nanopore. Contrary to SBS, it is therefore also able to identify modified and unnatural nucleotides such that the readout of data encoded using an expanded molecular alphabet is possible. , …”

“…Apart from DNA, other organic molecules have been recently proposed. , Two interesting examples are the use of peptide sequences for data storage, as reported by Ng et al and the use of urethanes as reported by Dahlhauser et al Unfortunately in both these cases, reading required the use of mass spectroscopy, with the consequent limitation in terms of costs and speed. Recent advances in nanopore-based readout of short peptide sequences may speed up developments in this area …”

Molecular Encryption and Steganography Using Mixtures of Simultaneously Sequenced, Sequence-Defined Oligourethanes

“…These macromolecules, often referred to as digital polymers, store information at the molecular level in the form of a defined and absolute monomer sequence (i.e., primary structure). − Encoding information at the molecular level can be used to surmount some drawbacks of conventional storage devices, such as durability, longevity, and excessive spatial occupation . Such macromolecular information storage is now well established with artificial DNA biopolymers, which have been shown to store and retrieve significant amounts of information. − Two strengths of using DNA for information storage are the ability to replicate and retrieve data, as well as the ability to exploit rapid advances in sequencing, such as Next-Gen methods , and nanopore technology. − Alongside DNA, advances in the synthesis and sequencing of abiotic SDPs have improved the information storage capabilities of these macromolecular systems, − with commensurate advances toward this goal seen with multicomponent reactions − and small molecule strategies. − However, significant advances are still needed to rival the effective storage capacity of nucleic acids using SDPs, let alone silicon-based data storage. , …”