KR-20260065496-A - High-Density Nucleic Acid Synthesis Substrate and Method for Manufacturing the Same

KR20260065496AKR 20260065496 AKR20260065496 AKR 20260065496AKR-20260065496-A

Abstract

The present invention relates to a high-density nucleic acid synthesis substrate and a method for manufacturing the same. The synthesis substrate of the present invention may include a plate-shaped main body, a plurality of synthesis sections penetrating the main body and having an inlet and an outlet, into which a porous glass bead (Controlled Pore Glass, CPG) is inserted, and a stopper that prevents the porous glass bead from leaking out from the outlet of the synthesis section. The present invention provides a new synthesis substrate that can simultaneously achieve the high integration of conventional 2D synthesis and the high yield of 3D synthesis, allows for the recovery of individual nucleic acids by separating porous glass beads after synthesis, and possesses low cost, high precision, and chemical resistance.

Inventors

방두희
김준형
김하은

Assignees

연세대학교 산학협력단

Dates

Publication Date: 20260508
Application Date: 20250912
Priority Date: 20241030

Claims (10)

Plate-shaped main body; A plurality of synthesis sections arranged at predetermined intervals along a plurality of rows and columns in the main body, each having an inlet on one side and an outlet on the other side respectively, formed in a column shape penetrating the main body, wherein nucleic acid supplied from the inlet combines with a porous glass bead (Controlled Pore Glass, CPG) inserted therein; and A high-density nucleic acid synthesis substrate comprising: a plurality of catch portions that shield a portion of the outlet on the opposite side of the main body portion to prevent the porous glass beads inserted inside the synthesis portion from flowing out through the outlet portion.
In paragraph 1, The above synthesis unit is, A high-density nucleic acid synthesis substrate characterized by being formed in a tapered shape in which the diameter decreases from the inlet to the outlet.
In paragraph 1, The diameter of the inlet of the above-mentioned synthesis part is, A high-density nucleic acid synthesis substrate characterized by having an average diameter of 1.2 times or more and 2.5 times or less than the average diameter of the porous glass beads.
In paragraph 1, The opening gap that is not shielded at the outlet of the composite part, where the above-mentioned catch part is, A high-density nucleic acid synthesis substrate characterized by having an average diameter of 0.6 times or less of the porous glass beads.
Step of preparing resin raw materials; A step of manufacturing a substrate having one of the shapes of claims 1 to 4 by 3D printing using the above resin raw material, comprising a plate-shaped main body, a plurality of composite parts formed to penetrate one side and the other side of the main body, and a catch portion at the end of the other side of the composite part; and A method for manufacturing a high-density nucleic acid synthesis substrate comprising the step of washing and post-curing the substrate.
In paragraph 5, The step of manufacturing the above substrate is, A method for manufacturing a high-density nucleic acid synthesis substrate characterized by manufacturing one side of the main body portion of the above substrate in contact with a glass plate.
In paragraph 6, The step of manufacturing the above substrate is, A method for manufacturing a high-density nucleic acid synthesis substrate characterized by treating the surface of the above glass plate with corona discharge.
In paragraph 5, The step of manufacturing the above substrate is, A method for manufacturing a high-density nucleic acid synthesis substrate characterized by the fact that the light irradiated above penetrates a PET film mounted on the bottom of a vat storing the resin raw material to selectively cure the resin raw material.
In paragraph 5, The step of preparing the above resin raw material is, A method for manufacturing a high-density nucleic acid synthesis substrate characterized by the above resin raw material being a UV absorber mixed with a UV-curable resin.
In paragraph 5, The step of cleaning and post-curing the above substrate is, A method for manufacturing a high-density nucleic acid synthesis substrate characterized by including the step of washing the substrate using ethanol or isopropyl alcohol.

Description

High-Density Nucleic Acid Synthesis Substrate and Method for Manufacturing the Same The present invention relates to a high-density nucleic acid synthesis substrate capable of simultaneously realizing high-yield characteristics of three-dimensional synthesis using porous glass beads and high-density characteristics of two-dimensional synthesis performed on the surface of a synthesis substrate, and a method for manufacturing the same. In general, microarray technology is widely used for the large-scale analysis and synthesis of nucleic acids, and can be broadly classified into analytical microarrays and synthetic microarrays. Analytical microarrays utilize a method that induces hybridization by applying fluorescently labeled samples to nucleic acids immobilized on a substrate surface, and then detects the binding status via a fluorescent signal using a microscope. While this method has been applied in gene expression profiling and diagnostic fields, its usage is currently on a downward trend due to limitations such as the instability of the immobilized nucleic acids and low signal reproducibility. Synthetic microarrays are a method of directly recovering nucleic acids synthesized on a substrate for use as a library, or attaching biotin to synthesized nucleic acids and then selectively recovering hybridized target sequences using magnetic beads coated with streptavidin. This method can be applied not only to diagnosis and profiling but also to nucleic acid library synthesis. The fundamental direction of development in microarray technology is to increase sequence density to enable the synthesis of more types of nucleic acid sequences within a limited substrate area. However, as sequence density increases, the area where individual sequences are synthesized has shrunk, resulting in a synthesis yield per sequence that is only at the femtomolar (fmol) level. Therefore, a PCR amplification process is essential for the actual utilization of synthesized nucleic acids. However, to perform PCR amplification, the synthesized nucleic acid must include a primer binding sequence, and there is a constraint that the nucleic acid length must be at least 60 nucleotides (nt) for stable amplification. This makes it difficult to synthesize short sequences and entails problems caused by amplification bias during the PCR process. Therefore, a new synthesis technique is required that achieves high directivity while increasing the amount of sequence synthesis to reduce dependence on PCR. The aforementioned background technology is technical information that the inventor possessed or acquired during the process of deriving the embodiments of the present invention, and it cannot be considered as prior art disclosed to the general public prior to the filing of the embodiments of the present invention. FIG. 1 shows a perspective view of a high-density nucleic acid synthesis substrate according to one embodiment of the present invention. FIGS. 2 and FIGS. 3 illustrate a perspective view and an enlarged view of a high-density nucleic acid synthesis substrate according to one embodiment of the present invention. FIG. 4 illustrates a side view of a high-density nucleic acid synthesis substrate according to one embodiment of the present invention. FIG. 5 illustrates a flowchart of a method for manufacturing a high-density nucleic acid synthesis substrate according to one embodiment of the present invention. FIG. 6 illustrates a photograph of a method for manufacturing a high-density nucleic acid synthesis substrate according to one embodiment of the present invention. FIG. 8 shows a photograph of a high-density nucleic acid synthesis substrate manufactured by a method for manufacturing a high-density nucleic acid synthesis substrate according to one embodiment of the present invention. The present invention will become clear from the embodiments described below in detail together with the accompanying drawings. However, the present invention is not limited to the embodiments disclosed below but may be implemented in various different forms. These embodiments are provided merely to ensure that the disclosure of the present invention is complete and to fully inform those skilled in the art of the scope of the invention, and the present invention is defined only by the scope of the claims. Meanwhile, the terms used in this specification are for describing the embodiments and are not intended to limit the present invention. Throughout this specification, the singular form includes the plural form unless specifically stated otherwise in the text. Throughout this specification, the terms “comprises” and/or “comprising” as used mean that the mentioned components, steps, actions and/or elements do not exclude the presence or addition of one or more other components, steps, actions and/or elements, and that, unless specifically stated otherwise, they do not exclude other components but may include additional components. Additionally, terms such as “