De novo synthesized gene libraries

Patent metadata

Field	Value
Publication number	US-2016303535-A1
Application number	US-201615187721-A
Country	US
Kind code	A1
Filing date	Jun 20, 2016
Priority date	Aug 5, 2013
Publication date	Oct 20, 2016
Grant date	—

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

1 . A device for nucleic acid assembly, comprising:
a) a first plate, the first plate comprising:
a base layer of the first plate; and
one or more wells, wherein each well of the one or more wells comprises:
a side wall that extends vertically from the base layer;
an interior region of the well; 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

b) a second plate, wherein the second plate comprises a plurality of clusters, wherein each cluster comprises a plurality of loci, wherein each locus comprises a physically distinct region 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 the first plate is silicon or plastic.

3 . The device of claim 1 , wherein the second plate is silicon.

4 . The device of claim 1 , wherein the interior region of the well has a width from 0.8 to 2.0 mm.

5 . The device of claim 4 , wherein the interior region of the well has a width of 1.15 mm.

6 . The device of claim 1 , wherein the side wall has a length from 0.1 to 0.6 mm.

7 . The device of claim 6 , wherein the side wall has a length of 0.45 mm.

8 . The device of claim 1 , wherein each well of the one or more wells comprises an aqueous liquid.

9 . The device of claim 8 , wherein the aqueous liquid has a surface tension from 12 to 80 millinewtons per meter.

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

11 . 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.

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

13 . The device of claim 12 , wherein the silane is methoxyethoxyundecyltrichlorosilane, methacryloxypropyltrimethoxysilane, ethyltrimethoxysilane, propyltrimethoxysilane, trifluoropropyltrimethoxysilane, 3-(2-aminoethy)aminopropyltrimethoxysilane, p-tolyltrimethoxysilane, cyanoethyltrimethoxysilane, aminopropyltriethoxysilane, acetoxypropyltrimethoxysilane, phenyltrimethoxysilane, chloropropyltrimethoxysilane, mercaptopropyltrimethoxysilane, trichloro-octodecyl-silane, methyltrimethoxysilane, butyl-aldehyde-trimethoxysilane, heptadecafluorodecyltrimethoxysilane, or glycidoxypropyltrimethoxysilane.

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

15 . The device of claim 14 , wherein the fluorosilane is (tridecafluorotetrahydrooctyl)-triethoxysilane.

16 . The device of claim 1 , wherein each locus of the plurality of loci comprises a microchannel.

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

Assignees

• Twist Bioscience Corp

Inventors

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

Classifications

• B01J2219/00596

  Solid-phase processes · CPC title

• B01J2219/00317

  Microwell devices, i.e. having large numbers of wells · CPC title

• C12Q1/6806

  Preparing nucleic acids for analysis, e.g. for polymerase chain reaction [PCR] assay (C12Q1/6804 takes precedence) · CPC title

• B01J2219/00709

  Type of synthesis · CPC title

• C40B50/14 Primary

  Solid phase synthesis, i.e. wherein one or more library building blocks are bound to a solid support during library creation; Particular methods of cleavage from the solid support · CPC title