Polymer sheets for sequencing applications

Abstract

Embodiments of the present application relate to patterned polymer sheets and processes to prepare the same for sequencing applications. In particular, flexible micro- and nano-patterned polymer sheets are prepared and used as a template surface in sequencing reaction and new polish-free methods of forming isolated hydrogel plugs in nanowells are described.

What is claimed is: 
     
         1 . A process for preparing a polymer sheet for sequencing applications, comprising:
 providing a substrate with a surface;   depositing a layer of a polymer composition onto the surface of the substrate, wherein the polymer composition comprises functional groups for grafting oligonucleotides;   forming the polymer composition into a polymer sheet on the surface; and   removing said polymer sheet from the surface of the substrate.   
     
     
         2 . The process of  claim 1 , wherein the polymer composition comprises a hydrogel. 
     
     
         3 . The process of  claim 1  or  2 , wherein the polymer composition comprises poly(N-(5-azidoacetamidylpentyl) acrylamide-co-acrylamide) (PAZAM). 
     
     
         4 . The process of any one of  claims 1  to  3 , wherein the forming of the polymer sheet comprises dehydrating the polymer composition. 
     
     
         5 . The process of  claim 4 , wherein the dehydrating is performed at an elevated temperature. 
     
     
         6 . The process of  claim 5 , wherein the dehydrating occurs at about 60° C. 
     
     
         7 . The process of any one of  claims 1  to  6 , wherein the surface of the substrate comprises micro-scale or nano-scale patterns. 
     
     
         8 . The process of  claim 7 , wherein the micro-scale or nano-scale patterns comprise channels, trenches, posts, wells, or combinations thereof. 
     
     
         9 . The process of  claim 7  or  8 , wherein an imprint of the micro-scale or nano-scale patterns of the patterned surface is transferred to the polymer sheet to form a patterned polymer sheet. 
     
     
         10 . A process for preparing a substrate surface for sequencing applications, comprising:
 providing a polymer sheet comprising a first plurality of functional groups;   contacting said polymer sheet with a surface of a substrate, wherein said surface comprises a second plurality of functional groups;   covalently bonding the first plurality of functional groups of the polymer sheet to the second plurality of functional groups of the surface.   
     
     
         11 . The process of  claim 10 , further comprising grafting oligonucleotides on the substrate surface by reacting functionalized oligonucleotides with the first plurality of functional groups of the polymer sheet. 
     
     
         12 . The process of  claim 10  or  11 , wherein the polymer sheet is a patterned polymer sheet comprising a plurality of micro-scale or nano-scale patterned channels, trenches, posts, wells, or combinations thereof. 
     
     
         13 . The process of any one of  claims 10  to  12 , wherein the polymer sheet comprises a plurality of nano-scale patterned posts or wells. 
     
     
         14 . The process of any one of  claims 10  to  13 , wherein the polymer sheet comprises a dehydrated hydrogel. 
     
     
         15 . The process of  claims 14 , wherein the polymer sheet is rehydrated prior to grafting oligonucleotides. 
     
     
         16 . The process of any one of  claims 10  to  15 , wherein the first plurality of functional groups are selected from C 8-14  cycloalkenes, 8 to 14 membered heterocycloalkenes, C 8-14  cycloalkynes, 8 to 14 membered heterocycloalkynes, alkynyl, vinyl, halo, azido, amino, amido, epoxy, glycidyl, carboxyl, hydrazonyl, hydrazinyl, hydroxy, tetrazolyl, tetrazinyl, nitrile oxide, nitrene, nitrone, oxo-amino, or thiol, or optionally substituted variants and combinations thereof. 
     
     
         17 . The process of  claim 16 , wherein the first plurality of functional groups are selected from azido, alkynyl, amino, carboxyl, epoxy, glycidyl, halo, or tetrazinyl, or optionally substituted variants and combinations thereof. 
     
     
         18 . The process of  claim 17 , wherein the first plurality of functional groups comprise azido. 
     
     
         19 . The process of any one of  claims 10  to  18 , wherein the polymer sheet comprises poly(N-(5-azidoacetamidylpentyl) acrylamide-co-acrylamide) (PAZAM). 
     
     
         20 . The process of any one of  claims 10  to  16 , wherein the second plurality of functional groups are selected from vinyl, acryloyl, alkenyl, alkynyl, C 8-14  cycloalkenes, 8 to 14 membered heterocycloalkenes, C 8-14  cycloalkynes, 8 to 14 membered heterocycloalkynes, nitrene, aldehyde, hydrazinyl, glycidyl ether, epoxy, amino, carbene, isocyanate or maleimide, or optionally substituted variants and combinations thereof. 
     
     
         21 . The process of  claim 20 , wherein the second plurality of functional groups are selected from alkynyl, acryloyl, C 8-14  cycloalkenes, alkynyl, glycidyl ether, epoxy, or optionally substituted variants and combinations thereof. 
     
     
         22 . The process of  claim 21 , wherein the second plurality of functional groups comprises optionally substituted norbornene. 
     
     
         23 . A process for preparing a patterned substrate surface for sequencing applications, comprising:
 providing a substrate with a patterned surface, wherein said surface comprises a plurality of micro-scale or nano-scale patterned wells;   depositing a polymer composition onto the patterned surface to form a first polymer layer, wherein the polymer composition fills the micro-scale or nano-scale patterned wells; and   separating the first polymer layer, wherein the polymer composition is isolated in the micro-scale or nano-scale patterned wells of the patterned surface.   
     
     
         24 . The process of  claim 23 , further comprising dehydrating the first polymer layer prior to the separating of the first polymer layer. 
     
     
         25 . The process of  claim 23  or  24 , wherein the polymer composition comprises a hydrogel. 
     
     
         26 . The process of any one of  claims 23  to  25 , wherein the polymer composition comprises a first plurality of functional groups selected from selected from vinyl, acryloyl, alkenyl, alkynyl, C 8-14  cycloalkenes, 8 to 14 membered heterocycloalkenes, C 8-14  cycloalkynes, 8 to 14 membered heterocycloalkynes, nitrene, aldehyde, hydrazinyl, glycidyl ether, epoxy, amino, carbene, isocyanate or maleimide, or optionally substituted variants and combinations thereof. 
     
     
         27 . The process of  claim 26 , wherein the first plurality of functional groups are selected from azido, alkynyl, amino, carboxyl, epoxy, glycidyl, halo, or tetrazinyl, or optionally substituted variants and combinations thereof. 
     
     
         28 . The process of  claim 27 , wherein the first plurality of functional groups comprise azido. 
     
     
         29 . The process of any one of  claims 23  to  28 , wherein the polymer composition comprises poly(N-(5-azidoacetamidylpentyl) acrylamide-co-acrylamide) (PAZAM). 
     
     
         30 . The process of any one of  claim 23  to  29 , further comprising laminating a second polymer layer directly on top of the patterned surface after separating the first polymer layer. 
     
     
         31 . The process of  claim 30 , wherein the second polymer layer comprises a second plurality of functional groups selected from vinyl, acryloyl, alkenyl, alkynyl, C 8-14  cycloalkenes, 8 to 14 membered heterocycloalkenes, C 8-14  cycloalkynes, 8 to 14 membered heterocycloalkynes, nitrene, aldehyde, hydrazinyl, glycidyl ether, epoxy, amino, carbene, isocyanate or maleimide, or optionally substituted variants and combinations thereof. 
     
     
         32 . The process of  claim 31 , wherein the second plurality of functional groups are selected from alkynyl, acryloyl, C 8-14  cycloalkenes, alkynyl, glycidyl ether, epoxy, or optionally substituted variants and combinations thereof. 
     
     
         33 . The process of  claim 32 , wherein the second plurality of functional groups comprises optionally substituted norbornene. 
     
     
         34 . The process of any one of  claims 30  to  33 , further comprising covalently bonding the first plurality of functional groups of the polymer composition to the second plurality of functional groups of the second polymer layer. 
     
     
         35 . The process of  claim 24 , wherein the polymer composition is rehydrated before reacting with the second plurality of functional groups. 
     
     
         36 . The process of  claim 34  or  35 , further comprising separating the second polymer layer from the patterned surface, wherein the polymer composition is removed from the micro-scale or nano-scale patterned wells of the patterned surface to form patterned polymer posts on the second polymer layer. 
     
     
         37 . The process of  claim 36 , wherein the polymer composition is dehydrated before separating the second polymer layer from the patterned surface. 
     
     
         38 . The process of any one of  claims 30  to  37 , wherein the second polymer layer is stretchable. 
     
     
         39 . The process of  claim 38 , wherein the second polymer layer comprises polydimethylsiloxane (PDMS), poly(methyl methacrylate) (PMMA), polyurethanes, polyethers, polycarbonates, polyvinyls, polyimides, or combinations and copolymers thereof. 
     
     
         40 . The process of  claim 37 , further comprising grafting oligonucleotides on the micro-scale or nano-scale patterned polymer posts of the second polymer layer. 
     
     
         41 . An automated roll-to-roll process for sequencing applications, comprising:
 spooling a roll of patterned polymer sheet prepared by the process of  claim 38 ;   preparing the patterned polymer sheet for sequencing by treating the polymer sheet with sequencing reagents;   sequencing a sample on the treated patterned polymer sheet; and   respooling the patterned polymer sheet after the completion of one sequencing cycle.   
     
     
         42 . The automated roll-to-roll process of  claim 41 , further comprising stretching the polymer sheet during imaging in the sequencing cycle. 
     
     
         43 . The automated roll-to-roll process of  claim 41 , wherein the patterned stretchable polymer sheet comprises polydimethylsiloxane (PDMS), poly(methyl methacrylate) (PMMA), polyurethanes, polyethers, polycarbonates, polyvinyls, polyimides, or combinations and copolymers thereof. 
     
     
         44 . A polymer sheet for sequencing applications prepared by any of the process of  claims 1  to  9 . 
     
     
         45 . A substrate surface for sequencing applications prepared by any of the process of  claims 10  to  40 . 
     
     
         46 . An automated process for sequencing applications, comprising:
 providing a belt comprising the patterned polymer sheet prepared by the process of  claim 38 ;   preparing the patterned polymer sheet for sequencing by treating the polymer sheet with sequencing reagents from a fluid delivery device; and   sequencing a sample on the treated patterned polymer sheet, wherein the belt passes the fluid delivery device for each cycle of the sequencing application.