De novo synthesized gene libraries
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.
1. A device for synthesis of nucleic acids, the device comprising:
a plate;
a plurality of clusters, wherein each cluster is located on a top surface of the plate, wherein a horizontal cross-sectional shape of each cluster is defined by a plurality of loci, wherein each cluster has a net surface energy that is higher than a surface energy for a region of the top surface of the plate adjacent to and surrounding each cluster, wherein the plurality of loci located within each cluster comprise 50 to 500 loci, and wherein each locus has a total width of up to 500 um; and
a plurality of posts located within each locus, wherein each post extends from the top surface of the plate, has a horizontal cross-section that is rectangular or circular in shape, is separated from a neighboring post, has a width of up to 1 um, and has a height from the top surface of the plate of up to 1 um, wherein a ratio of the width to the height of each post is from 0.1 to 1, wherein each post comprises a plurality of oligonucleotides bound thereto, wherein each of the oligonucleotides comprises at least 80 nucleotides in length, and wherein the plurality of posts provide for rapid exchange of chemical exposure during de novo synthesis of oligonucleotides.
2. The device of claim 1 , wherein the ratio of the width to the height of each post is from 0.4 to 0.8.
3. The device of claim 1 , wherein each of the posts has rounded edges.
4. The device of claim 1 , wherein each locus has a total width of 1 um to 500 um.
5. The device of claim 1 , wherein each locus has a total width of 1 um to 100 um.
6. The device of claim 1 , wherein a total number of the loci on the surface comprises 700,000 loci.
7. The device of claim 1 , wherein the width of each locus is up to 300 times larger than the width of each of the posts.
8. The device of claim 1 , wherein each cluster provides for a water contact angle that is up to 90 degrees greater than a water contact angle of the region of the top surface adjacent to and surrounding each cluster.
9. The device of claim 1 , wherein each cluster provides for a water contact angle that is up to 20 degrees greater than a water contact angle of the region of the top surface adjacent to and surrounding each cluster.
10. The device of claim 1 , wherein each cluster has a square or round shape.
11. The device of claim 1 , wherein each cluster has a width of 0.5 mm to 1.5 mm.
12. The device of claim 1 , wherein each locus comprises a first plurality of molecules, and wherein the first plurality of molecules are each bound to the top surface of the plate and comprise a reactive group capable of binding to a nucleoside.
13. The device of claim 12 , wherein the first plurality of molecules comprises a silane or a siloxane.
14. The device of claim 12 , wherein the first plurality of molecules comprises 11-acetoxyundecyltriethoxysilane, n-decyltriethoxysilane, (3-aminopropyl)trimethoxysilane, or (3-aminopropyl)triethoxysilane or N-(3-triethoxysilylpropyl)-4-hydroxybutyramide.
15. The device of claim 12 , wherein a region surrounding each locus comprises a second plurality of molecules, wherein the second plurality of molecules are each bound to the top surface of the plate and lack a reactive group capable of binding to a nucleoside.
16. The device of claim 15 , wherein the second plurality of molecules comprises a fluorosilane.
17. The device of claim 16 , wherein the fluorosilane is (tridecafluorotetrahydrooctyl)-triethoxysilane.
18. The device of claim 1 , wherein each locus comprises a plurality of first molecules, wherein each first molecule is bound to the top surface of the plate and comprises a reactive group capable of binding to a nucleoside, and wherein a region adjacent to and surrounding each locus comprises a plurality of second molecules, wherein each second molecule is bound to the top surface of the plate and lacks the reactive group capable of binding to the nucleoside, and wherein each locus has a higher surface energy than the region adjacent to and surrounding each locus.
19. The device of claim 1 , wherein the plate comprises nylon, nitrocellulose, or polypropylene.
20. The device of claim 1 , wherein the plate comprises glass, silica, silicon, polytetraflouroethylene, polypropylene, polystyrene, polycarbonate, gold, platinum, agarose, dextran, polyacrylamides, or polydimethylsiloxane (PDMS).
21. The device of claim 1 , wherein the plate comprises a silicon layer.
22. The device of claim 1 , wherein each of the oligonucleotides comprises at least 100 nucleotides in length.