Systems and methods for storing and reading nucleic acid-based data with error protection

Abstract

The systems, devices, and methods described herein provide scalable methods for writing data to and reading data from nucleic acid molecules. The present disclosure covers four primary areas of interest: (1) accurately and quickly reading information stored in nucleic acid molecules, (2) partitioning data to efficiently encode data in nucleic acid molecules, (3) error protection and correction when encoding data in nucleic acid molecules, and (4) data structures to provide efficient access to information stored in nucleic acid molecules.

1 . A method for reading information stored in nucleic acid sequences, the method comprising:
 obtaining a pool of identifier nucleic acid molecules that store digital information from a string of symbols of length L, wherein each individual identifier nucleic acid molecule comprises a plurality of component nucleic acid molecules and corresponds to a symbol value and a symbol position in the string of symbols, and wherein the pool of identifier nucleic acid molecules correspond to a subset of identifier nucleic acid sequences in an identifier library that is capable of encoding any string of symbols having length L;   reading an identifier nucleic acid molecule of the obtained identifier nucleic acid molecules to identify a read sequence corresponding to a portion of the identifier nucleic acid molecule;   identifying, based on the read sequence, a set of candidate identifier nucleic acid sequences each (i) corresponding to an entry in the identifier library and (ii) comprising a component nucleic acid molecule having a sequence that approximates or exactly matches the read sequence;   assigning, to each candidate identifier nucleic acid sequence, a score representative of how similar the respective candidate identifier nucleic acid sequence is to the identifier nucleic acid molecule; and   selecting, based on the scores, one of the candidate identifier nucleic acid sequences as a selected sequence.   
     
     
         2 . The method of  claim 1 , further comprising using the identifier library to map the selected sequence to one of the symbol positions and one of the symbol values within the string of symbols. 
     
     
         3 . The method of  claim 2 , further comprising determining additional symbol positions and symbol values within the string of symbols by using the identifier library to map additional selected sequences corresponding to likely sequences of identifier nucleic acid molecules in the pool. 
     
     
         4 . The method of  claim 1 , wherein reading the identifier nucleic acid molecule comprises sequencing at least a portion of the identifier nucleic acid molecule by at least one selected from the group of: chemical sequencing, chain termination sequencing, shotgun sequencing, bridge PCR sequencing, single-molecule real-time sequencing, ion semiconductor sequencing, pyrosequencing, sequencing by synthesis, combinatorial probe anchor synthesis sequencing, sequencing by ligation, nanopore sequencing, nanochannel sequencing, massively parallel signature sequencing, Polony sequencing, DNA nanoball sequencing, single molecule fluorescent sequencing, tunneling current sequencing, sequencing by hybridization, mass spectrometry sequencing, microfluidic sequencing, transmission electron microscopy sequencing, RNA polymerase sequencing, and in vitro virus sequencing. 
     
     
         5 . The method of  claim 4 , wherein the sequencing comprises:
 applying an electric field to an electrolytic solution and at least one nanopore channel;   translocating the identifier nucleic acid molecule through the at least one nanopore channel; and   measuring impedance in the at least one nanopore channel, wherein the component nucleic acid sequences each have a corresponding unique impedance signature along the length of the sequence.   
     
     
         6 . The method of  claim 5 , wherein sequencing at least one component nucleic acid sequence in the identifier nucleic acid molecule comprises comparing measured impedance values to the unique impedance signature. 
     
     
         7 . The method of  claim 5 , wherein the at least one nanopore channel is formed from alpha-hemolysin (αHL) or mycobacterium smegmatis porin A (MspA). 
     
     
         8 . The method of  claim 5 , wherein the at least one nanopore channel is formed within a solid-state membrane. 
     
     
         9 . The method of  claim 5 , further comprising, prior to reading an identifier nucleic acid molecule, ligating the at least one identifier nucleic acid molecule to a second identifier nucleic acid molecule. 
     
     
         10 . The method of  claim 5 , further comprising, prior to reading the identifier nucleic acid molecule, degrading one strand of the at least one identifier nucleic acid molecule. 
     
     
         11 . The method of  claim 10 , wherein a strand-specific exonuclease is used to selectively degrade one strand of the at least one identifier nucleic acid molecule. 
     
     
         12 . The method of  claim 5 , wherein the electric field generates a differential potential greater than 100 mV across the at least one nanopore channel, and wherein translocating the at least one identifier nucleic acid molecule occurs at a rate greater than 1,000 bases per second. 
     
     
         13 . The method of  claim 5 , further comprising, before translocating, binding an agent to the at least one identifier nucleic acid molecule, wherein the agent is associated with an agent signature in the measure impedance. 
     
     
         14 . The method of  claim 13 , wherein at least one unique impedance signature comprises the agent signature, and wherein determining at least one component nucleic acid sequence in the identifier nucleic acid molecule comprises comparing measured impedance values to said at least one unique impedance signature. 
     
     
         15 . The method of  claim 13 , wherein a presence of the agent on the at least one nucleic acid molecule allows for a first maximum translocation rate that achieves a desired level of accuracy that is faster than a second maximum translocation rate that achieves the desired level of accuracy in the absence of the agent on the at least one nucleic acid molecule. 
     
     
         16 . The method of  claim 13 , wherein binding the agent to the at least one identifier nucleic acid molecule comprises using an enzyme, and wherein binding the agent to the at least one identifier nucleic acid molecule occurs at a known location on a component nucleic acid molecule such that the agent signature at the known location results in a known shift in impedance value during translocation. 
     
     
         17 . The method of  claim 16 , wherein the agent is a base analogue or a modified base, and the enzyme is a polymerase, wherein the polymerase incorporates the base analogue or the modified base in the at least one identifier nucleic acid molecule during replication. 
     
     
         18 . The method of  claim 13 , further comprising binding each agent of a plurality of agents at known locations on the plurality of component nucleic acid molecules, wherein the plurality of agents and known locations of each agent comprise an agent signature. 
     
     
         19 . The method of  claim 16 , wherein the enzyme is methyltransferase. 
     
     
         20 . The method of  claim 1 , wherein the obtained identifier nucleic acid molecules are encoded with a minimum number of base difference from one another such that each identifier nucleic acid molecule is associated with a reading error tolerance. 
     
     
         21 . The method of  claim 20 , wherein the reading error tolerance permits faster reading of the identifier nucleic acid molecule. 
     
     
         22 . The method of  claim 1 , further comprising:
 determining, based on mapping the selected sequence to one of the symbol positions and one of the symbol values within the string of symbols, a decoded string of symbols;   calculating a hash of a portion of the decoded string of symbols;   comparing the calculated hash to an original hash associated with a corresponding portion of the string of symbols; and   verifying, based on the comparison, whether the portion of the decoded string of symbols matches the portion of the string of symbols.   
     
     
         23 . The method of  claim 22 , further comprising:
 determining the portion of the decoded string of symbols does not match the portion of the string of symbols;   selecting, based on the scores, a second candidate identifier nucleic acid sequence as the selected sequence; and   using the identifier library to map the selected sequence to one of the symbol positions and one of the symbol values within the string of symbols.   
     
     
         24 . The method of  claim 22 , wherein the hash of the portion of the decoded string of symbols is calculated using at least one of: MD5, SHA-224, SHA-256, SHA-384, SHA-512, SHA-512/224, or SHA-512/256. 
     
     
         25 . The method of  claim 1 , further comprising:
 computing a sample size estimate for the pool of identifier nucleic acid molecules; and   sampling the pool of identifier nucleic acid molecules, based on the sample size estimate, to obtain the identifier nucleic acid molecule.   
     
     
         26 . The method of  claim 1 , wherein each identifier nucleic acid molecule in the pool of nucleic acid molecules comprises M component nucleic acid molecules corresponding to M layers. 
     
     
         27 . The method of  claim 26 , wherein reading the identifier nucleic acid molecule includes reading N of the M layers. 
     
     
         28 . The method of  claim 1 , further comprising replicating the identifier nucleic acid molecule such that the identifier nucleic acid molecule comprises modified bases. 
     
     
         29 . The method of  claim 1 , wherein the score is a distance metric representative of a degree of similarity between the respective candidate identifier nucleic acid sequence and the identifier nucleic acid molecule. 
     
     
         30 - 92 . (canceled) 
     
     
         93 . The method of  claim 1 , wherein a presence or absence of an identifier nucleic acid molecule in the pool is representative of the symbol value of the corresponding respective symbol position within the string of symbols.