[학위논문]핵산 나노 구조체의 대량 생산 및 선택적 장시간 방출을 위한 핵산 기반의 신규 물질 개발 Development of nucleic acid-based novel materials for mass production and selective long-term release of nucleic acid nanostructure원문보기
핵산은 왓슨-크릭 (Watson-Crick) 염기 쌍에 기초한 빌딩 블록 재료로서 주목을 받고 있다. 핵산 기반 나노-, 마이크로기술에서, 4가지 유형의 단위체 (Adenosine, Thymine (Uracil), Guanine, Cytosine)는 2D 기반의 junction 나노구조체부터 ...
핵산은 왓슨-크릭 (Watson-Crick) 염기 쌍에 기초한 빌딩 블록 재료로서 주목을 받고 있다. 핵산 기반 나노-, 마이크로기술에서, 4가지 유형의 단위체 (Adenosine, Thymine (Uracil), Guanine, Cytosine)는 2D 기반의 junction 나노구조체부터 3D 기반의 origami까지 컴퓨터 프로그래밍 되었다. 또한, 최근에는 수많은 기능성 핵산을 함유하는 중합체 입자 및 하이드로젤이 생의학 응용을 위해 연구되었다. 생체 적합성 및 생체 내 서열을 모방하는 능력에 기초하여, 핵산 기반 물질은 약물 전달을 위한 약물 운반체, 사전에 병원체를 검출하기 위한 바이오 센서 및 특정 유전자 서열을 사용한 면역학적 치료제로서 연구되어왔다. 그러나, 이러한 개발에도 불구하고, 핵산의 통상적인 합성 방법 (예를 들어, PCR, 클로닝)은 비용 및 수율 측면에서 한계를 보여 왔으며, 이는 핵산 기반 재료의 생체 의학적 응용을 위한 상용화에 장벽으로 작용하고 있다. 우리는 이러한 장벽을 극복하기 위해 롤링 서클 복제법을 이용하였다. 롤링 서클 복제법은 값비싼 thermal cycler를 사용하지 않으며 반복적인 핵산 기반 구조체의 대량 생산을 위한 비용 측면에 효율적인 방법의 하나다. 이 논문에서는 첫 번째, RCA를 기반으로 한 DNA 나노구조체의 대량 증폭과 선택적 분리 프로세스를 보였으며 두 번째, RNA particle을 bio-ink로 이용하여 약물 모델로서 안정성과 전달 효율이 높은 마이크로구조체 제작에 대한 프린트 시스템을 연구하였다. 우리는 대량 증폭된 핵산을 기반으로 한 새로운 유형의 물질 개발과 약물 모델로서의 정교한 방출 제어에 대한 본 연구가 핵산의 의학적 응용 분야에 잠재적인 기여를 할 것으로 기대한다.
핵산은 왓슨-크릭 (Watson-Crick) 염기 쌍에 기초한 빌딩 블록 재료로서 주목을 받고 있다. 핵산 기반 나노-, 마이크로기술에서, 4가지 유형의 단위체 (Adenosine, Thymine (Uracil), Guanine, Cytosine)는 2D 기반의 junction 나노구조체부터 3D 기반의 origami까지 컴퓨터 프로그래밍 되었다. 또한, 최근에는 수많은 기능성 핵산을 함유하는 중합체 입자 및 하이드로젤이 생의학 응용을 위해 연구되었다. 생체 적합성 및 생체 내 서열을 모방하는 능력에 기초하여, 핵산 기반 물질은 약물 전달을 위한 약물 운반체, 사전에 병원체를 검출하기 위한 바이오 센서 및 특정 유전자 서열을 사용한 면역학적 치료제로서 연구되어왔다. 그러나, 이러한 개발에도 불구하고, 핵산의 통상적인 합성 방법 (예를 들어, PCR, 클로닝)은 비용 및 수율 측면에서 한계를 보여 왔으며, 이는 핵산 기반 재료의 생체 의학적 응용을 위한 상용화에 장벽으로 작용하고 있다. 우리는 이러한 장벽을 극복하기 위해 롤링 서클 복제법을 이용하였다. 롤링 서클 복제법은 값비싼 thermal cycler를 사용하지 않으며 반복적인 핵산 기반 구조체의 대량 생산을 위한 비용 측면에 효율적인 방법의 하나다. 이 논문에서는 첫 번째, RCA를 기반으로 한 DNA 나노구조체의 대량 증폭과 선택적 분리 프로세스를 보였으며 두 번째, RNA particle을 bio-ink로 이용하여 약물 모델로서 안정성과 전달 효율이 높은 마이크로구조체 제작에 대한 프린트 시스템을 연구하였다. 우리는 대량 증폭된 핵산을 기반으로 한 새로운 유형의 물질 개발과 약물 모델로서의 정교한 방출 제어에 대한 본 연구가 핵산의 의학적 응용 분야에 잠재적인 기여를 할 것으로 기대한다.
From simple 2-dimensional (2D) structures to highly complex 3-dimensional (3D) structures, DNA is widely used as a building block material. Despite this advancement, mass production of DNA nanostructures was a challenge due to the high cost that accompanies the preparation of diverse DNA strands. Ro...
From simple 2-dimensional (2D) structures to highly complex 3-dimensional (3D) structures, DNA is widely used as a building block material. Despite this advancement, mass production of DNA nanostructures was a challenge due to the high cost that accompanies the preparation of diverse DNA strands. Rolling circle amplification (RCA) is one of the cost-effective methods for the mass production of repetitive DNA structures without using an expensive laboratory thermal cycler. Here, we report a DNA hydrogel containing various single-stranded DNA nanostructures (Y, X, Tetrahedron) in a one-pot system, prepared by RCA, and their selective separation with various restriction enzymes. Each DNA nanostructure in the DNA hydrogel contained distinctive restriction sites that were responsive to different kinds of restriction enzymes, achieving selective separation of Y-, X-, and Tetrahedron-shaped DNA structures. This study has the potential to contribute towards DNA-based biomedical applications by employing mass production and precisely controlled release of DNA nanostructures.The RNA nanotechnology has been advanced as functional materials via enzymatic replication and rational programming. However, the delivery efficiency of functional RNA is reduced due to the degradation by RNase and the negative charge of the RNA's backbone. Here, to overcome these drawbacks, we propose an RNA particle-based microstructure in which RNA particles are densely networked via a 3D bio-printing system. This disk shapes microstructure was showed the long-term release of functional RNA and high stability to RNase. Furthermore, to increase delivery efficiency as a carrier, the RNA disk was encapsulated with DNA hydrogel or chitosan/hyaluronic acid layer. This universal our platform is expected to provide high-efficient loading and delivery system of functional RNA.The nucleic acid has attracted attention as a building block material based on Watson-Crick base pairs. In the nucleic acid-based nano-, micro-technology, four types of monomer (Adenosine, Thymine (Uracil), Guanine, Cytosine) were computer programmed from 2D-based junction nanostructures to 3D-based complex origami. Furthermore, recently, the polymeric particle and hydrogel containing countless functionalized nucleic acid were invented for biomedical applications. Based on their biocompatibility and the ability to mimicking in vivo sequences, nucleic acid-based materials have been studied as drug carriers for drug delivery, biosensors to detect pathogens in advance and immunological therapeutics using specific gene sequences. However, despite these developments, conventional synthetic methods (e.g., PCR, cloning) of nucleic acid have been shown limitations in terms of cost and yield, and it is a barrier to the commercialization of nucleic acid-based materials for biomedical application. To overcome this barrier, we applicate the rolling circle replication to this study. rolling circle replication is one of the cost-effective methods for the mass production of repetitive nucleic acid-based structures without using an expensive laboratory thermal cycler. In this thesis, we first demonstrated rolling circle amplification (RCA)-based mass amplification and the selective separation process of DNA nanostructures. Second, we used the RNA particles as bio-inks to produce microstructures with high stability and delivery efficiency as drug models. We expect that this study about mass amplified nucleic acid-based new types of material and rationally controlled release, as a drug model, potentially contributes to biomedical application fields.
From simple 2-dimensional (2D) structures to highly complex 3-dimensional (3D) structures, DNA is widely used as a building block material. Despite this advancement, mass production of DNA nanostructures was a challenge due to the high cost that accompanies the preparation of diverse DNA strands. Rolling circle amplification (RCA) is one of the cost-effective methods for the mass production of repetitive DNA structures without using an expensive laboratory thermal cycler. Here, we report a DNA hydrogel containing various single-stranded DNA nanostructures (Y, X, Tetrahedron) in a one-pot system, prepared by RCA, and their selective separation with various restriction enzymes. Each DNA nanostructure in the DNA hydrogel contained distinctive restriction sites that were responsive to different kinds of restriction enzymes, achieving selective separation of Y-, X-, and Tetrahedron-shaped DNA structures. This study has the potential to contribute towards DNA-based biomedical applications by employing mass production and precisely controlled release of DNA nanostructures.The RNA nanotechnology has been advanced as functional materials via enzymatic replication and rational programming. However, the delivery efficiency of functional RNA is reduced due to the degradation by RNase and the negative charge of the RNA's backbone. Here, to overcome these drawbacks, we propose an RNA particle-based microstructure in which RNA particles are densely networked via a 3D bio-printing system. This disk shapes microstructure was showed the long-term release of functional RNA and high stability to RNase. Furthermore, to increase delivery efficiency as a carrier, the RNA disk was encapsulated with DNA hydrogel or chitosan/hyaluronic acid layer. This universal our platform is expected to provide high-efficient loading and delivery system of functional RNA.The nucleic acid has attracted attention as a building block material based on Watson-Crick base pairs. In the nucleic acid-based nano-, micro-technology, four types of monomer (Adenosine, Thymine (Uracil), Guanine, Cytosine) were computer programmed from 2D-based junction nanostructures to 3D-based complex origami. Furthermore, recently, the polymeric particle and hydrogel containing countless functionalized nucleic acid were invented for biomedical applications. Based on their biocompatibility and the ability to mimicking in vivo sequences, nucleic acid-based materials have been studied as drug carriers for drug delivery, biosensors to detect pathogens in advance and immunological therapeutics using specific gene sequences. However, despite these developments, conventional synthetic methods (e.g., PCR, cloning) of nucleic acid have been shown limitations in terms of cost and yield, and it is a barrier to the commercialization of nucleic acid-based materials for biomedical application. To overcome this barrier, we applicate the rolling circle replication to this study. rolling circle replication is one of the cost-effective methods for the mass production of repetitive nucleic acid-based structures without using an expensive laboratory thermal cycler. In this thesis, we first demonstrated rolling circle amplification (RCA)-based mass amplification and the selective separation process of DNA nanostructures. Second, we used the RNA particles as bio-inks to produce microstructures with high stability and delivery efficiency as drug models. We expect that this study about mass amplified nucleic acid-based new types of material and rationally controlled release, as a drug model, potentially contributes to biomedical application fields.
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