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US-12618061-B2 - Methods of gene assembly using DNAzymes and use in DNA data storage

US12618061B2US 12618061 B2US12618061 B2US 12618061B2US-12618061-B2

Abstract

Building DNA strands at a high rate that are suitable for data storage. Methods include using DNAzyme and utilizing libraries of pre-prepared oligos. A system for the DNA strand synthesis includes: a DNA symbol library comprising a number of single strand oligo symbols; a DNA linker library comprising a first set of single strand oligo linkers and a second set of single strand oligo linkers; and a DNAzyme library comprising a number of DNAzymes. An S1 end of a first DNAzyme is adapted to join the S1 end of a symbol and an S2 end of the first DNAzyme is adapted to join an S2 end of a first linker, and an S1 end of a second DNAzyme is adapted to join an S1 end of a second linker and an S2 end of the second DNAzyme is adapted to join an S2 end of the symbol.

Inventors

  • Gemma Roselle MENDONSA
  • Walter R. Eppler

Assignees

  • SEAGATE TECHNOLOGY LLC

Dates

Publication Date
20260505
Application Date
20220729

Claims (16)

  1. 1 . A system for DNA synthesis, comprising: a DNA symbol library comprising a number of single strand oligo symbols, each symbol having an activated 3′-phosphorimidazolide S1 end and a 5′-hydroxyl S2 end; a DNA linker library comprising a first set of single strand oligo linkers each having a 3′-phosphorimidazolide S1 end and a 5′-hydroxyl S2 end, and a second set of single strand oligo linkers each having an activated 3′-phosphorimidazolide S1 end and a 5′-hydroxyl S2 end; and a DNAzyme library comprising a number of DNA ligating DNAzymes, each DNA ligating DNAzyme having a 3′-phosphorimidazolide complementary S1 end and a 5′-hydroxyl complementary S2 end; wherein each of the linkers has a sequence specificity for the DNA ligating DNAzymes, and one or both of the S1 end and the S2 end of each of the linkers hybridizes to a complementary one of the DNA ligating DNAzymes; and wherein: the S1 end of a first DNA ligating DNAzyme joins the S1 end of a symbol and the S2 end of the first DNA ligating DNAzyme joins the S2 end of a first linker, and the S1 end of a second DNA ligating DNAzyme joins the S1 end of a second linker and the S2 end of the second DNA ligating DNAzyme joins the S2 end of the symbol.
  2. 2 . The system of claim 1 , wherein the symbols comprise a base section having four to eight nucleotides, and a 3′-phosphorimidazolide S1 end section and a 5′-hydroxyl S2 end section each having six to ten nucleotides, independently.
  3. 3 . The system of claim 2 , wherein the symbols comprise two joined nibbles, each nibble having a base sub-section and an end section.
  4. 4 . The system of claim 1 , wherein the linkers comprise six to twenty nucleotides.
  5. 5 . The system of claim 1 , wherein the DNA ligating DNAzymes are E47 DNAzymes.
  6. 6 . A system for DNA synthesis, comprising: a first DNA symbol library comprising a number of single strand oligo first nibble symbols, each first nibble symbol having a base section and a 3′-phosphorimidazolide S1 end; a second DNA symbol library comprising a number of single strand oligo second nibble symbols, each second nibble symbol having a base section and a 5′-hydroxyl S2 end; a DNA linker library comprising a first set of single strand oligo linkers each having a 3′-phosphorimidazolide S1 end and a 5′-hydroxyl S2 end, and a second set of single strand oligo linkers each having a 3′-phosphorimidazolide S1 end and a 5′-hydroxyl S2 end; and a DNAzyme library comprising a number of DNA ligating DNAzymes, each DNA ligating DNAzyme having a 3′-phosphorimidazolide complementary S1 end and a 5′-hydroxyl complementary S2 end; wherein: wherein each of the linkers has a sequence specificity for the DNA ligating DNAzymes, and one or both of the S1 end and the S2 end of each of the linkers hybridizes to a complementary one of the DNA ligating DNAzymes; the base section of a first nibble symbol joins the base section of a second nibble symbol; the S1 end of a first DNA ligating DNAzyme joins the S1 end of one of the first nibble symbols and the S2 end of the first DNA ligating DNAzyme joins the S2 end of a first linker, and the S1 end of a second DNA ligating DNAzyme joins the S1 end of a second linker and the S2 end of the second DNA ligating DNAzyme joins the S2 end of one of the second nibble symbols.
  7. 7 . The system of claim 6 , wherein the first nibble symbols and the second nibble symbols have combined base sections having four to eight nucleotides, the first nibble symbol has a 3′-phosphorimidazolide S1 end section having six to ten nucleotides, and the second nibble symbol has a 5′-hydroxyl S2 end section each having six to ten nucleotides.
  8. 8 . The system of claim 6 , wherein the linkers comprise six to twenty nucleotides.
  9. 9 . The system of claim 6 , wherein, combined, the first DNA symbol library and the second DNA symbol library has 256 nibbles.
  10. 10 . A method of making a DNA strand, comprising: providing a DNA symbol library comprising a number of single strand DNA oligo symbols, each symbol having a 3′-phosphorimidazolide S1 end and a 5′-hydroxyl S2 end; providing a DNA linker library comprising a first set of single strand DNA oligo linkers having a 3′-phosphorimidazolide S1 end and a second set of single strand DNA oligo linkers having a 5′-hydroxyl S2 end, wherein each of the linkers has a sequence specificity for the DNA ligating DNAzymes, and one or both of the S1 end and the S2 end of each of the linkers hybridizes to a complementary one of the DNA ligating DNAzymes; providing a DNAzyme comprising a number of DNA ligating DNAzymes having a 3′-phosphorimidazolide complementary S1 end and a 5′-hydroxyl complementary S2 end; joining the S1 end of a first DNA ligating DNAzyme to the S1 end of a symbol and the S2 end of the first DNA ligating DNAzyme to the S2 end of a first linker; and joining the S1 end of a second DNA ligating DNAzyme to the S1 end of a second linker and the S2 end of the second DNA ligating DNAzyme to the S2 end of the symbol.
  11. 11 . The method of claim 10 , wherein: joining the S1 end of a first DNA ligating DNAzyme to the S1 end of a symbol and the S2 end of the first DNA ligating DNAzyme to the S2 end of a first linker; and joining the S1 end of a second DNA ligating DNAzyme to the S1 end of a second linker and the S2 end of the second DNA ligating DNAzyme to the S2 end of the symbol, are done simultaneously.
  12. 12 . The method of claim 10 , further comprising joining the DNA strand made in claim 10 with a second DNA strand.
  13. 13 . The method of claim 12 , wherein joining the DNA strand made in claim 10 with the second DNA strand is via an enzyme assembly.
  14. 14 . The method of claim 12 , wherein joining the DNA strand made in claim 10 with the second DNA strand is via another DNA ligating DNAzyme.
  15. 15 . The method of claim 1 , wherein the DNA ligating DNAzymes are E47 DNAzymes.
  16. 16 . The system of claim 1 , wherein the DNA ligating DNAzymes are E47 DNAzymes.

Description

BACKGROUND There is always a desire for more data storage and increased speed writing to and reading from that storage, as well as a desire for reduced cost for the data storage. DNA is an emerging technology for data storage. Current methods assert that a DNA strand or gene, to store 5 KB of data, can be written in 14 days. Comparatively, magnetic disk drives and magnetic tapes both can write 1 TByte in about an hour. A single DNA base pair location can store 2 bits; thus, 4000 Giga-base pairs would need to be stored in an hour to match the capabilities of a single disk drive or tape. Although current technology is believed to be capable of writing 15 base pairs an hour, there needs to be an 8 to 9 order of magnitude improvement in order for DNA data storage to be viable. In addition to the speed differential between magnetic disk drives and tapes and DNA data storage, magnetic media data storage is a mature technology, optimized in many ways including cost. New processes need to be developed to make DNA data storage economical. SUMMARY This disclosure is directed to methods of building DNA strands, or genes, at a high rate that are suitable for data storage. The methods include using DNAzyme and utilizing libraries of pre-prepared oligos that are combined to form the desired DNA gene, encoding the desired data. One particular implementation described herein is a system for making a DNA gene. One system comprises a DNA symbol library comprising a number of single strand oligo symbols, each symbol having an S1 end and an S2 end, a DNA linker library comprising a first set of single strand oligo linkers each having an S1 end and a second set of single strand oligo linkers each having an S2 end, and a DNAzyme library comprising a number of DNAzymes, each DNAzyme having an S1 end and an S2 end. The S1 end of a first DNAzyme is adapted to join the S1 end of a symbol and the S2 end of the first DNAzyme is adapted to join the S2 end of a first linker, and the S1 end of a second DNAzyme is adapted to join the S1 end of a second linker and the S2 end of the second DNAzyme is adapted to join the S2 end of the symbol. Another particular system described herein comprises a first DNA symbol library comprising a number of single strand oligo first nibble symbols, each nibble symbol having a base section and an S1 end, a second DNA symbol library comprising a number of single strand oligo second nibble symbols, each nibble symbol having a base section and an S2 end, a DNA linker library comprising a first set of single strand oligo linkers each having an S1 end and a second set of single strand oligo linkers each having an S2 end, and a DNAzyme library comprising a number of DNAzymes, each DNAzyme having an S1 end and an S2 end. The base section of a first nibble symbol is adapted to join the base section of a second nibble symbol, the S1 end of a first DNAzyme is adapted to join the S1 end of the first nibble symbol and the S2 end of the first DNAzyme is adapted to join the S2 end of a first linker, and the S1 end of a second DNAzyme is adapted to join the S1 end of a second linker and the S2 end of the second DNAzyme is adapted to join the S2 end of the second nibble symbol. Another particular implementation described herein is a method of making a DNA strand. The method comprises providing a DNA symbol library comprising a number of single strand DNA oligo symbols, each symbol having an S1 end and an S2 end, providing a DNA linker library comprising a first set of single strand DNA oligo linkers having an S1 end and a second set of single strand DNA oligo linkers having an S2 end, providing a DNAzyme comprising a number of DNAzymes having an S1 end and an S2 end, joining the S1 end of a first DNAzyme to the S1 end of a symbol and the S2 end of the first DNAzyme to the S2 end of a first linker, and joining the S1 end of a second DNAzyme to the S1 end of a second linker and the S2 end of the second DNAzyme to the S2 end of the symbol. Other systems and methods are also described herein. This Summary is provided to introduce a selection of concepts in a simplified form that are further described below in the Detailed Description. This Summary is not intended to identify key features or essential features of the claimed subject matter, nor is it intended to be used to limit the scope of the claimed subject matter. These and various other features and advantages will be apparent from a reading of the following detailed description. BRIEF DESCRIPTION OF THE DRAWING The described technology is best understood from the following Detailed Description describing various implementations read in connection with the accompanying drawing. FIG. 1 is a schematic rendering of a DNAzyme joining two oligos. FIG. 2A is a schematic rendering of a first linker oligo; FIG. 2B is a schematic rendering of a symbol oligo; FIG. 2C is s schematic rendering of a second linker oligo; FIG. 2D is a schematic rendering of a first DNAzyme; and FIG. 2E