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 the recording latency and synthesis cost of oligos using the four natural nucleotides remain high. Here, we describe an improved DNA-based storage system that uses an extended 11-letter molecular alphabet combining natural and chemically modified nucleotides. Our extended-alphabet molecular storage paradigm offers a nearly two-fold increase in storage density and potentially the same order of reduction in the recording time. Experimental results involving a library of 77 custom-designed hybrid sequences reveal that one can readily detect and discriminate different combinations and orders of monomers via MspA nanopores. Furthermore, a neural network architecture designed to classify raw current signals generated by Oxford Nanopore Technologies sequencing ensures an average accuracy exceeding 60%, which is 39 times higher than that of random guessing. Molecular dynamics simulations reveal that the majority of modified nucleotides do not induce dramatic disruption of the DNA double helix, making the extended alphabet system potentially compatible with PCR-based random access data retrieval. The methodologies proposed provide a forward path for new implementations of molecular recorders.

Emerging Approaches to DNA Data Storage: Challenges and Prospects

“…4,[42][43][44][45][46][47][48][49][50]36,14 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 7,51,52 or chemically modified nucleotides. 53 2.2. Storage and Degradation Issues.…”

“…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. 53,87 While nanopore sequencing improves upon several limitations of SBS for DNA data storage, as reviewed by Ceze et al, 12 two key constraints of the technology are its high error rate and the required sequence length. The high error rate of nanopore sequencing (∼10% per nt in the single read), 70,88 compared with the nearly negligible rate of errors introduced by SBS (∼0.5% per nt), 70 necessitates the clustering of sequence information, and thus, higher sequencing coverage and additional postprocessing of sequencing data.…”

“…Recent advances in nanopore-based readout of short peptide sequences 135 may speed up developments in this area. 53 3.3. DNA Nanotechnology for Molecular Computation.…”