De novo synthesized gene libraries

Patent metadata

Field	Value
Publication number	US-9403141-B2
Application number	US-201514885963-A
Country	US
Kind code	B2
Filing date	Oct 16, 2015
Priority date	Aug 5, 2013
Publication date	Aug 2, 2016
Grant date	Aug 2, 2016

Abstract

De novo synthesized large libraries of nucleic acids are provided herein with low error rates. Further, devices for the manufacturing of high-quality building blocks, such as oligonucleotides, are described herein. Longer nucleic acids can be synthesized in parallel using microfluidic assemblies. Further, methods herein allow for the fast construction of large libraries of long, high-quality genes. Devices for the manufacturing of large libraries of long and high-quality nucleic acids are further described herein.

First claim

What is claimed is:

1. A device for nucleic acid assembly, comprising:
(i) a first silicon plate, the first silicon plate comprising:
a base layer of the first silicon plate;
an insulator layer on top of the base layer, wherein the insulator layer comprises silicon dioxide; and
one or more wells, wherein each well of the one or more wells comprises:
a side wall that extends vertically from the insulator layer, wherein the side wall has a length from 0.1 to 0.6 mm;
an interior region of the well having a width from 0.8 to 2.0 mm; and
an exterior region of the well, wherein the exterior region of the well has a lower surface energy than the interior region of the well; and

(ii) a second silicon plate, wherein the second silicon plate comprises a plurality of clusters, wherein each cluster comprises a plurality of loci, wherein each locus comprises a physically distinct coating comprising molecules capable of binding to a nucleoside phosphoramidite, and wherein each cluster is vertically aligned to a single well of the one or more wells.

2. The device of claim 1 , wherein each well of the one or more wells comprises an aqueous liquid, wherein the aqueous liquid has a surface tension from 12 to 80 millinewtons per meter.

3. The device of claim 1 , wherein the width of the interior region of the well is 1.15 mm, and wherein the length of the side wall is 0.45 mm.

4. The device of claim 1 , wherein the device further comprises an enclosure, wherein the first silicon plate and second silicon plate are both within the enclosure.

5. The device of claim 1 , wherein each well of the one or more wells comprises a sealing means located at a distal end of the side wall of said well.

6. The device of claim 1 , wherein the interior region of each well of the one or more wells comprises a silane.

7. The device of claim 6 , wherein the silane is methoxyethoxyundecyltrichlorosilane, methacryloxypropyltrimethoxysilane, ethyltrimethoxysilane, propyltrimethoxysilane, trifluoropropyltrimethoxysilane, 3-(2-aminoethyl)aminopropyltrimethoxysilane, p-tolyltrimethoxysilane, cyanoethyltrimethoxysilane, aminopropyltriethoxysilane, acetoxypropyltrimethoxysilane, phenyltrimethoxysilane, chloropropyltrimethoxysilane, mercaptopropyltrimethoxysilane, trichloro-octodecyl-silane, methyltrimethoxysilane, butyl-aldehyde-trimethoxysilane, heptadecafluorodecyltrimethoxysilane, or glycidoxypropyltrimethoxysilane.

8. The device of claim 1 , wherein the interior region of each well of the one or more wells comprises an aminosilane.

9. The device of claim 1 , wherein the exterior region of each well of the one or more wells comprises a fluorosilane.

10. The device of claim 9 , wherein the fluorosilane is (tridecafluorotetrahydrooctyl)-triethoxysilane.

11. The device of claim 1 , wherein each cluster of the plurality of clusters of the second silicon plate has a lower surface energy than the interior region of each well the plurality of wells of the first silicon plate.

12. The device of claim 1 , where each locus of the plurality of loci comprises a microchannel.

Assignees

• Twist Bioscience Corp

Inventors

• Banyai William
• Peck Bill James
• Fernandez Andres
• Chen Siyuan
• Indermuhle Pierre

Classifications

• B01J2219/00637

  by coating it with another layer · CPC title

• B01J2219/00497

  Features relating to the solid phase supports · CPC title

• C40B40/06

  Libraries containing nucleotides or polynucleotides, or derivatives thereof · CPC title

• B01J2219/00722

  Nucleotides · CPC title

• B01J2219/00619

  using hydrophilic or hydrophobic regions · CPC title