Dynamic information storage method based on DNA (deoxyribonucleic acid) aggregate and application of dynamic information storage method

Abstract

The invention belongs to the technical field of DNA information storage, and relates to a dynamic information storage method based on a DNA condensation body and application thereof, the method comprises the following steps: coding file information into a DNA base sequence according to a coding rule, adding a cohesive end at a 3'end to obtain two complementary single-stranded DNAs, and synthesizing a DNA double-stranded structure with the cohesive end through co-annealing; the method comprises the following steps: synthesizing an X-type DNA structure by using four DNA single chains with cohesive ends at 3'ends through co-annealing; mixing the DNA double-chain structure with the X-type DNA structure, and incubating at room temperature in a salt ion solution to obtain a DNA aggregate containing DNA information; carrying out target information amplification and sequencing on the solution of the DNA condensation body containing the DNA information by utilizing an amplification primer, and decoding to obtain file information so as to realize reading of the file information; the DNA condensation body is used as a liquid-phase storage medium, and information reading and dynamic operation can be achieved.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of DNA information storage, and particularly relates to a dynamic information storage method based on DNA condensates and application thereof. BACKGROUND

[0002] With the advent of the "digital era", the amount of data created globally per year is showing explosive growth, and the carrying capacity of traditional magnetic and optical storage media will soon reach the physical limit. DNA-based data storage technology has the advantages of high storage density, long life and low energy consumption, which is conducive to saving storage energy and promoting the development of big data storage. The typical process of DNA information storage includes information encoding, DNA synthesis, information storage, information reading and information decoding.

[0003] At present, the method based on DNA information storage mainly focuses on realizing long-term and stable "cold storage" application, and significant progress has been made in improving storage density and storage stability. For example, high stability and high density information storage can be realized by encapsulating DNA information through media such as silicon dioxide and hydrogel. However, the above scheme mainly faces the archived data which does not need to be frequently accessed. When information needs to be read or processed, complex pretreatment steps often need to be introduced, such as adding specific chemical reagents, changing temperature or pH value to destroy the encapsulation structure, and realizing the release of DNA information. These pretreatment operations not only significantly increase the time cost and operation complexity of information processing, but also may introduce the risk of information loss and damage in the process of DNA release, thereby affecting the accuracy of information reading.

[0004] In addition, in other reported systems for dynamic processing of information, the combination between the storage medium and the DNA information molecule often depends on non-specific interaction. In order to realize the accurate processing of specific information, additional screening markers such as fluorescence and optical markers need to be added in such systems. This additional screening marker and screening process not only greatly reduces the operation simplicity of the storage system, but also affects the fidelity of information processing and the integration of the system.

[0005] Therefore, it is urgent to develop a new type of DNA information storage system which has the ability of high-density storage of "cold storage" data, and can realize the immediate reading and accurate dynamic operation of information which needs to be frequently processed. SUMMARY

[0006] The present application aims to overcome the deficiencies in the prior art and provide a dynamic information storage method based on DNA condensates and application thereof.

[0007] To achieve the purpose of the present application, the technical scheme is as follows.

[0008] A dynamic information storage reading method based on DNA condensate, comprising the following steps: S1. According to the encoding rule, encode the file information into DNA base sequence, and add sticky ends at the 3' end of the DNA base sequence to obtain two complementary single-stranded DNA, and synthesize DNA double-stranded structure with sticky ends by co-annealing; S2. Four designed DNA single strands with 3' end sticky ends are used to synthesize X-type DNA structure by co-annealing; S3. Mix the synthesized DNA double-stranded structure with sticky ends with X-type DNA structure, incubate at room temperature in a solution containing salt ions to obtain DNA condensate containing DNA information, i.e. store DNA information into DNA condensate; S4. Take the sample from the solution containing DNA condensate, use the designed amplification primer to amplify and sequence the target information, decode to obtain the file information, so as to realize the reading of file information.

[0009] Further, the format of the file information includes text, picture, video and audio.

[0010] Further, the file information is converted into binary data form according to the general conversion rule, and according to the encoding rule: four bases A, T, C, G represent 00, 01, 10, 11 respectively; and then converted into DNA sequence information. Further, the co-annealing conditions are: 95℃ 5min; 95℃-25℃ (-1℃ / min); 4℃ ∞.

[0011] Further, the molar ratio of the X-type DNA structure mixed with the DNA double-stranded structure is: 1:1-1:10.

[0012] Further, the salt ion is any one of sodium ion, potassium ion and magnesium ion.

[0013] Further, the concentration of the salt ion is 12.5mM-500mM.

[0014] Further, the pH value of the solution is 7.8-8.2.

[0015] Further, the incubation time is 5-60min, and the shaking speed is 0-330rpm.

[0016] A method for specifically erasing DNA information from DNA condensates involves adding an erasing strand to the DNA condensate containing DNA information prepared using steps S1 to S3 of the dynamic information storage and retrieval method described in claim 1, followed by incubation at room temperature to specifically erase the DNA information; wherein the erasing strand is designed based on information from the DNA base sequence and the X-type DNA structural sequence.

[0017] Furthermore, all of the DNA double-stranded structures share the same region that binds to the X-type DNA structure.

[0018] Furthermore, the erasure strand is a single-stranded DNA designed based on the DNA base sequence and the X-type DNA structural sequence, which is complementary to the sticky ends of the DNA double-stranded structure.

[0019] A method for repeatedly erasing and writing DNA information in DNA condensates includes the following steps: S121. The method described in claim 9 is used to erase DNA information, and the erased DNA information strands are washed away from the supernatant by centrifugation. S122. Add the buffer solution for forming DNA aggregates, resuspend the precipitate left in step S121, and centrifuge again to wash away the remaining erased DNA information strands in the supernatant. S123. Add the DNA information strand and buffer solution for forming DNA aggregates back into the buffer solution, and incubate at room temperature to achieve the rewriting of DNA information. S124. Repeat steps S121-S123 to repeatedly erase and write DNA information.

[0020] A method for replacing DNA information includes the following steps: S131. The method according to claim 9 achieves the erasure of DNA information by centrifugation to wash away the erased DNA information strands in the supernatant; S132. Add the buffer solution for forming DNA aggregates, resuspend the precipitate left in step S131, and centrifuge again to wash away the remaining erased DNA information strands in the supernatant. S133. Add new DNA information strands and buffer solution for forming DNA aggregates, and incubate at room temperature to achieve the replacement of different DNA information.

[0021] Compared with the prior art, the present invention has the following significant advantages: (1) The present invention provides a DNA condensate as a liquid storage medium, which can realize frequent and convenient information reading and dynamic operation without destroying the encapsulation structure; (2) The DNA condensate storage method provided by the present invention can realize precise dynamic operation of information without the need for additional screening markers; (3) The binding between the storage medium and DNA information molecules is based on the principle of complementary base pairing, which has high programmability and specificity, ensuring the fidelity of information processing and the degree of system integration; (4) The present invention provides a dynamic storage medium based on DNA condensates that has the ability to store “cold storage” data in high density and the function of accurately and dynamically processing “hot storage” data. Attached Figure Description

[0022] Figure 1 This is a schematic diagram of the dynamic information storage method and its application described in this invention; Figure 2 A schematic diagram of the structural information strand of a double-stranded DNA with sticky ends; Figure 3 Confocal microscopy results of storing DNA information in DNA condensates; Figure 4 Figure (a) shows a schematic diagram of agarose gel electrophoresis for information reading; Figure (b) shows a Sanger sequencing diagram. Figure 5 Image of the results of fluorescence confocal microscopy for erasing specific information; Figure 6 A graph showing the changes in DNA information content during a repeatable information erasure-write operation. Detailed Implementation

[0023] All reagents used in this invention are commercially available. All DNA sequences were synthesized by Sangon Biotech (Shanghai) Co., Ltd.

[0024] According to the encoding rules, the text information is encoded into the corresponding DNA sequence information, which is then synthesized by Sangon Biotech (Shanghai) Co., Ltd.

[0025] Table 1. DNA sequence information encoded by text information

[0026] The primer sequence information required for reading and amplifying the target sequence was designed and synthesized by Sangon Biotech (Shanghai) Co., Ltd.

[0027] Table 2 DNA sequence information of amplification primers The DNA sequence information required for designing and synthesizing the X-type DNA structure was commissioned to Sangon Biotech (Shanghai) Co., Ltd.

[0028] Table 3. DNA sequence information required for synthesizing X-type DNA structures First, the two complementary single-stranded DNAs corresponding to the file information are co-annealed to synthesize a DNA double-stranded structure with sticky ends, which serves as the DNA information strand; then, the X-type DNA structure is co-annealed to synthesize the DNA double-stranded structure. The DNA information strand contains an ID region for specific recognition and amplification, such as... Figure 2 As shown, DNA strands containing target information are selectively amplified using amplification primers.

[0029] The prepared DNA double-stranded structure and X-type DNA structure were mixed in a reaction solution (10 mM Tris-HCl, 350 mM Na). + In a solution (pH 8.0), with final concentrations of 500 nM and 5 μM, incubation was performed at room temperature with shaking (330 rpm) for 0-30 min to achieve DNA information storage. Figure 3 As shown, DNA information can be stored in DNA condensates in just 5 minutes.

[0030] Take 1 μL of the DNA condensate solution containing the information and amplify the target information using the corresponding primers. For example... Figure 4 As shown, agarose gel electrophoresis and Sanger sequencing confirmed the correct reading of the information.

[0031] Based on the DNA sequence information and X-type DNA structural sequence information obtained in Example 1, a corresponding erasure strand was designed and synthesized by Sangon Biotech (Shanghai) Co., Ltd. The erasure strand is a single-stranded DNA. The erasure strand is completely complementary to the sticky end region of the DNA information strand. Figure 2 DNA information strands containing different information can specifically bind to different erasure strands. First, the erasure strand specifically recognizes and binds to the free regions in the sticky ends of the DNA information strand, and then competitively binds to the hybridization region of the DNA information strand and the X-type DNA structure, thereby separating the DNA information strand from the X-type DNA structure and releasing the DNA information strand from the DNA condensate.

[0032] Table 4. DNA sequence information of the strand to be erased for specific information erasure. Add the corresponding erasure strand to the DNA condensate solution containing mixed information, with a molar ratio of information strand to erasure strand of 1:10, and incubate at room temperature for 20 min to achieve specific erasure of DNA information. Figure 5 As shown, specific erasure of DNA information can be achieved within 20 minutes.

[0033] Based on the specific information erasure scheme in Example 2, the information is first erased.

[0034] For samples after the information erasure operation, centrifuge at 4℃ and 450xg for 30 min to wash away the deleted DNA information strands in the supernatant; Add the buffer solution for DNA aggregation (10mM Tris-HCl, 350mM Na) + (pH 8.0), the precipitate left after resuspending and centrifugation was washed again by centrifugation at 4℃ and 450xg for 30 min to remove the remaining deleted DNA information strands in the supernatant; Re-add the DNA information strand (final concentration 500 nM) and the buffer for forming DNA aggregates (10 mM Tris-HCl, 350 mM Na) + Incubate at room temperature (pH 8.0) with shaking (330 rpm) for 15 min to rewrite DNA information; Repeating the above steps will achieve a repeatable information erase-write cycle. For example... Figure 6 As shown, during the six erase-write cycles, the concentration of DNA information stored in the DNA condensate exhibits regular fluctuations, demonstrating that repeatable erase-write cycles can be achieved in the dynamic storage system of DNA condensates.

[0035] The preferred embodiments of the present application have been described above with reference to the accompanying drawings, but this does not limit the scope of the claims of the present application. Any modifications, equivalent substitutions, and improvements made by those skilled in the art without departing from the scope and substance of the embodiments of the present application shall be within the scope of the claims of the present application.

Claims

1. A dynamic information storage reading method based on DNA condensates, characterized by: The method comprises the following steps: S1, according to the encoding rule, encoding the file information into DNA base sequence, and adding sticky ends to the 3' end of the DNA base sequence to obtain two complementary single-stranded DNAs, and synthesizing a DNA double-stranded structure with sticky ends through co-annealing; S2, four designed DNA single strands with 3' end sticky ends are used to synthesize an X-type DNA structure through co-annealing; S3, the synthesized DNA double-stranded structure with sticky ends is mixed with the X-type DNA structure, and incubated at room temperature in a solution containing salt ions to obtain a DNA condensate containing DNA information, that is, the DNA information can be stored in the DNA condensate; S4, a sample is taken from the solution containing the DNA condensate, and the designed amplification primer is used for amplification and sequencing of the target information, and the file information is decoded to realize reading of the file information.

2. A dynamic information storage reading method based on DNA condensates according to claim 1, characterized in that: The format of the file information includes text, picture, video and audio.

3. The dynamic information storage reading method based on DNA condensates according to claim 1, characterized in that: The co-annealing condition is 95℃ for 5min; 95℃ to 25℃ (-1℃ / min); 4℃ ∞.

4. The dynamic information storage reading method based on DNA condensates according to claim 1, characterized in that: The molar ratio of the X-type DNA structure to the DNA double-stranded structure is 1:1 to 10:

1.

5. The dynamic information storage reading method based on DNA condensates according to claim 1, characterized in that: The salt ion is any one of sodium ion, potassium ion and magnesium ion.

6. The dynamic information storage reading method based on DNA condensates according to claim 1, characterized in that: The concentration of the salt ion is 12.5mM to 500mM.

7. The dynamic information storage reading method based on DNA condensates according to claim 1, characterized in that: The pH value of the solution is 7.8 to 8.

2.

8. The dynamic information storage reading method based on DNA condensates according to claim 1, characterized in that: The incubation time is 5 to 60min, and the shaking speed is 0 to 330rpm.

9. A method for specifically erasing the information of DNA in a DNA condensate, characterized by: An erasing chain is added to the DNA condensate containing DNA information prepared in steps S1 to S3 of the dynamic information storage and reading method of claim 1, and incubated at room temperature, so that the DNA information can be specifically erased; wherein: the erasing chain is designed according to the information of the DNA base sequence and the X-type DNA structure sequence.

10. The method of claim 9, wherein the DNA condensates are specific to DNA. The DNA double-stranded structure shares the same region combined with the X-type DNA structure.

11. The method of claim 9, wherein the method is specific for erasing the information of the DNA in the DNA condensates. The erasing chain is a single-stranded DNA designed according to the DNA base sequence and the X-type DNA structure sequence, which can be complementary to the sticky end of the DNA double-stranded structure.

12. A method of repeatedly erasing and writing information in DNA condensates, comprising: The method comprises the following steps: S121, the method of claim 9 is used to realize the erasure of the DNA information, and the supernatant is washed away by centrifugation; S122, the buffer solution for forming the DNA condensate is added, the precipitate left in step S121 is resuspended, and the supernatant is washed away by centrifugation again; S123, the DNA information chain and the buffer solution for forming the DNA condensate are added again, and incubated at room temperature to realize the re-writing of the DNA information; S124, steps S121-S123 are repeated, and the erased and written DNA information can be repeated.

13. A method of replacing DNA information, characterized by: The method comprises the following steps: S131, the method of claim 9 is used to realize the erasure of the DNA information, and the supernatant is washed away by centrifugation; S132, the buffer solution for forming the DNA condensate is added, the precipitate left in step S131 is resuspended, and the supernatant is washed away by centrifugation again; S133, add new DNA information strand and buffer solution of DNA condensate formation, incubate at room temperature to achieve different DNA information replacement.

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