CHEMICAL METHODS FOR NUCLEIC ACID-BASED DATA STORAGE
The present disclosure discloses methods and systems for encoding digital information in nucleic acid (e.g., deoxyribonucleic acid) molecules without base-by-base synthesis, by encoding bit-value information in the presence or absence of unique nucleic acid sequences within a pool, comprising specifying each bit location in a bit-stream with a unique nucleic sequence and specifying the bit value at that location by the presence or absence of the corresponding unique nucleic acid sequence in the pool. Also disclosed are chemical methods for generating unique nucleic acid sequences using combinatorial genomic strategies (e.g., assembly of multiple nucleic acid sequences or enzymatic-based editing of nucleic acid sequences).
1 - 63 . (canceled)
64 . A method for writing information into a nucleic acid sequence, comprising:
(a) providing a device comprising a plurality of partitions;
(b) determining a string of symbols to represent said information;
(c) constructing, on at least one partition, a plurality of components, wherein each individual component of said plurality of components is a nucleic acid molecule having a nucleic acid sequence;
(d) altering the temperature in the at least one partition; and
(e) chemically linking together two or more components of said plurality of components, thereby generating one or more identifiers of an identifier library, wherein each identifier of said one or more identifiers comprises two or more components.
65 . The method of claim 64 , wherein an individual identifier of said one or more identifiers corresponds to an individual symbol in said string of symbols.
66 . The method of claim 65 , wherein one symbol value at each position of said string of symbols is represented by the absence of a distinct identifier in the identifier library.
67 . The method of claim 64 , wherein said two or more components are assembled in a fixed order.
68 . The method of claim 64 , wherein said two or more components are assembled with one or more partitioning components disposed between two components from different layers of said two or more layers.
69 . The method of claim 64 , wherein said plurality of nucleic acid sequences stores metadata of said information or conceals said information.
70 . The method of claim 64 , further comprising:
combining two or more identifier libraries; and
tagging each identifier library of said two or more identifier libraries with a distinct barcode.
71 . The method of claim 64 , wherein each individual identifier in said identifier library comprises a distinct barcode.
72 . The method of claim 71 , wherein said distinct barcode of each individual identifier has a minimum hamming distance from the distinct barcode of other individual identifiers.
73 . The method of claim 64 , wherein chemically linking comprises hybridizing two single-stranded nucleic acid molecules.
74 . The method of claim 73 , wherein hybridizing comprises annealing two single-stranded nucleic acid molecules.
75 . The method of claim 64 , wherein chemically linking comprises ligating together two or more components of said plurality of components using a reagent.
76 . The method of claim 75 , wherein said reagent is a T4 ligase, a T7 ligase, a T3 ligase, Taq ligase, Chlorella virus ligase, Thermococcus sp. strain 9° N ligase, or an E. coli ligase.
77 . The method of claim 76 , further comprising inactivating said ligases using a buffer containing EDTA or guanidine thiocyanate.
78 . The method of claim 75 , wherein said reagent comprises polyethylene glycol (PEG), dimethyl sulfoxide (DMSO), 1,2-Propanediol (1,2-Prd) glycerol, or polysorbate 20.
79 . The method of claim 75 , wherein said reagent further comprises glycerol molecules.
80 . The method of claim 64 , wherein chemically linking in (e) comprises using overlap-extension polymerase chain reaction (PCR).
81 . The method of claim 64 , further comprising dehydrating said identifier library by dehydrating each individual identifier of at least said subset of said one or more identifiers.
82 . The method of claim 64 , further comprising amplifying said one or more identifiers with PCR or linear amplification.
83 . The method of claim 82 , wherein said PCR has at least 10 cycles.