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Kafe 바로가기국가/구분 | United States(US) Patent 등록 |
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국제특허분류(IPC7판) |
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출원번호 | US-0135434 (2016-04-21) |
등록번호 | US-9981239 (2018-05-29) |
발명자 / 주소 |
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출원인 / 주소 |
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대리인 / 주소 |
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인용정보 | 피인용 횟수 : 0 인용 특허 : 471 |
Devices and methods for de novo synthesis of large and highly accurate libraries of oligonucleic acids are provided herein. Devices include structures having a main channel and microchannels, where the microchannels have a high surface area to volume ratio. Devices disclosed herein provide for de no
Devices and methods for de novo synthesis of large and highly accurate libraries of oligonucleic acids are provided herein. Devices include structures having a main channel and microchannels, where the microchannels have a high surface area to volume ratio. Devices disclosed herein provide for de novo synthesis of oligonucleic acids having a low error rate.
1. A device for synthesizing oligonucleotides, comprising: a plate;a plurality of main channels, wherein each main channel extends vertically into the plate from an opening on a top side of the plate, and wherein the plurality of main channels comprises more than 250 main channels; andfor each main
1. A device for synthesizing oligonucleotides, comprising: a plate;a plurality of main channels, wherein each main channel extends vertically into the plate from an opening on a top side of the plate, and wherein the plurality of main channels comprises more than 250 main channels; andfor each main channel, a plurality of microchannels connected thereto, wherein each microchannel of the plurality of microchannels extends vertically from an opening on a bottom side of the plate to said main channel, and wherein each microchannel of the plurality of microchannels has a surface area to volume ratio of greater than 0.2 um−1. 2. The device of claim 1, wherein the surface area to volume ratio provides for rapid exchange of chemical exposure during de novo synthesis of oligonucleotides. 3. The device of claim 1, wherein the surface area to volume ratio is about 0.4 um−1. 4. The device of claim 1, wherein each microchannel of the plurality of microchannels has a surface area greater than 10,000 um2. 5. The device of claim 4, wherein each microchannel of the plurality of microchannels has a surface area of about 13,000 um2. 6. The device of claim 1, wherein the plurality of microchannels comprises 50 to 500 microchannels. 7. The device of claim 1, wherein a ratio of width to depth of a narrowest segment of each microchannel is from 0.5 to 0.01. 8. The device of claim 1, wherein each microchannel of the plurality of microchannels has a total width of 30 um to 100 um. 9. The device of claim 1, wherein each microchannel of the plurality of microchannels has a depth of 10 um to 500 um. 10. The device of claim 1, wherein each main channel has a width from 0.5 mm to 2 mm. 11. The device of claim 1, wherein the plurality of main channels comprises about 10,000 main channels. 12. The device of claim 1, further comprising a plurality of first molecules, wherein each first molecule is bound to an interior surface of the plurality of microchannels and comprises a reactive group capable of binding to a nucleoside phosphoramidite. 13. The device of claim 12, wherein each first molecule is a silane or a siloxane. 14. The device of claim 13, wherein each first molecule is an aminosilane. 15. The device of claim 12, wherein each first molecule is selected from the group consisting of 11-acetoxyundecyltriethoxysilane, n-decyltriethoxysilane, (3-aminopropyl)trimethoxysilane, (3-aminopropyl)triethoxysilane, glycidyloxypropyl/trimethoxysilane, and N-(3-triethoxysilylpropyl)-4-hydroxybutyramide. 16. The device of claim 12, further comprising a plurality of second molecules, wherein each second molecule is bound to an interior surface of the main channel and lacks the reactive group capable of binding to the nucleoside phosphoramidite. 17. The device of claim 16, wherein each second molecule is a fluorosilane. 18. The device of claim 17, wherein the fluorosilane is (tridecafluorotetrahydrooctyl)-triethoxysilane. 19. The device of claim 16, wherein the first molecule has a higher surface energy than the second molecule. 20. The device of claim 1, wherein the plate is a silicon plate. 21. A device for synthesizing oligonucleotides, comprising: a silicon plate;a plurality of main channels, wherein the plurality of main channels comprises more than 250 main channels, wherein each main channel extends vertically into the silicon plate from an opening on a top side of the silicon plate, and wherein each main channel has a width of 0.5 mm to 2 mm; andfor each main channel, a plurality of microchannels connected thereto, wherein each microchannel of the plurality of microchannels extends vertically from an opening on a bottom side of the silicon plate to said main channel, and wherein each microchannel of the plurality of microchannels comprises a total width that is less than 100 um, a microchannel surface area greater than 10,000 um2, and a maximum width for a narrowest segment of the microchannel of 10 um. 22. A method for de novo oligonucleotide synthesis, comprising: (a) providing predetermined sequences for a plurality of non-identical oligonucleotides;(b) providing the device of claim 1;(c) adding a droplet of fluid comprising an extension reaction reagent specific to a microchannel;(d) allowing sufficient time for an extension reaction step to occur; and(e) repeating steps (c) and (d) until the plurality of non-identical oligonucleotides are synthesized, wherein each oligonucleotide comprises at least 10 bases in length and is attached to an inside region of the microchannel. 23. The method of claim 22, wherein the synthesized non-identical oligonucleotides encode sequences with an aggregate error rate of less than 1 in 2000 bases compared to the predetermined sequences. 24. The method of claim 22, further comprising washing the plate with a washing reagent, and wherein washing removes greater than 95% of unincorporated extension reaction reagent. 25. The method of claim 24, wherein washing removes greater than 99% of unincorporated extension reaction reagent. 26. The method of claim 22, wherein the synthesized plurality of non-identical oligonucleotides collectively encode for at least 200 genes at least 1 kb in length. 27. The method of claim 22, further comprising: releasing the plurality of non-identical oligonucleotides from the plate; andsubjecting the plurality of non-identical oligonucleotides to a polymerase chain assembly reaction to assemble at least 200 genes. 28. The method of claim 27, wherein the at least 200 genes have an aggregate error rate of less than 1 in 2000 bases compared to the predetermined sequences without correcting errors.
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