Methods for Attaching Cellular Constituents to a Matrix

Abstract

The present invention relates to a method of covalently attaching a chromosome within a cell to a matrix including modifying a plurality of nucleotides within the chromosome to include a matrix attachment moiety wherein the chromosome contacts the matrix, and attaching the matrix attachment moiety of the plurality of nucleotides to the matrix, thereby attaching the chromosome to the matrix.

1 .- 29 . (canceled) 
     
     
         30 . A method of modeling structure of a chromosome in a cell comprising
 hybridizing to the chromosome a plurality of oligonucleotides, wherein the chromosome contacts a matrix within the cell, and   attaching the plurality of oligonucleotides to the matrix,   wherein the plurality of oligonucleotides attached to the matrix represents the structure of the chromosome in the cell.   
     
     
         31 . The method of  claim 30  wherein a matrix-forming material is introduced into the cell and forms the matrix contacting the chromosome. 
     
     
         32 . The method of  claim 30  wherein the matrix is expanded and additional oligonucleotides are hybridized to the chromosome and attached to the matrix. 
     
     
         33 . The method of  claim 30  wherein the matrix is repeatedly expanded and additional oligonucleotides are hybridized to the chromosome and attached to the matrix. 
     
     
         34 . The method of  claim 30  wherein the matrix-forming material is polymerized to form the matrix. 
     
     
         35 . The method of  claim 30  wherein the matrix-forming material is crosslinked to form the matrix. 
     
     
         36 . The method of  claim 30  further including removing the chromosome from the material, wherein the plurality of oligonucleotides attached to the matrix material represents the structure of the chromosome in the cell. 
     
     
         37 . The method of  claim 30  wherein the plurality of oligonucleotides include a matrix attachment moiety that is attached to the matrix. 
     
     
         38 . The method of  claim 30  wherein the plurality of oligonucleotides includes a matrix attachment moiety that is attached to the matrix and wherein the plurality of oligonucleotides includes DNA attachment moieties and the DNA attachment moieties are attached to chromosomal DNA. 
     
     
         39 . The method of  claim 30  wherein the plurality of oligonucleotides is chemically modified to include a matrix attachment moiety and the matrix attachment moiety is attached to the matrix. 
     
     
         40 . The method of  claim 30  wherein the plurality of oligonucleotides include a secondary probe hybridization site and a secondary probe including a matrix attachment moiety is hybridized to the secondary probe hybridization site and the matrix attachment moiety is attached to the matrix. 
     
     
         41 . The method of  claim 30  wherein the plurality of oligonucleotides includes a plurality of secondary probe hybridization sites and secondary probes including a matrix attachment moiety is hybridized to the plurality of secondary probe hybridization sites and the matrix attachment moiety is attached to the matrix. 
     
     
         42 . The method of  claim 30 , wherein the plurality of oligonucleotides comprises one or more detectable labels. 
     
     
         43 . The method of  claim 42 , further comprising detecting the detectable labels. 
     
     
         44 . The method of  claim 43 , wherein the one or more detectable labels comprises a fluorescent label and wherein the step of detecting the detectable labels comprises fluorescence microscopy, super-resolution microscopy, Stimulated Emission Depletion Microscopy (STED), structured Illumination Microscopy (SIM), and single-molecule super-resolution microscopy technologies selected from Photo-Activated Localization Microscopy (PALM), stochastic Optical Reconstruction Microscopy (STORM) and DNA-based Point Accumulation In Nanoscale Topography (DNA-PAINT). 
     
     
         45 . The method of  claim 44 , wherein prior to the step of detecting the detectable labels, the matrix is expanded. 
     
     
         46 . The method of  claim 40 , wherein one or more of the secondary probes comprises a detectable label. 
     
     
         47 . The method of  claim 46 , further comprising detecting the detectable label. 
     
     
         48 . The method of  claim 47 , wherein the detectable marker comprises a fluorescent label, and wherein the step of detecting the detectable label comprises one or more of fluorescence microscopy, super-resolution microscopy, Stimulated Emission Depletion Microscopy (STED), structured Illumination Microscopy (SIM), and single-molecule super-resolution microscopy technologies selected from Photo-Activated Localization Microscopy (PALM), stochastic Optical Reconstruction Microscopy (STORM) and DNA-based Point Accumulation In Nanoscale Topography (DNA-PAINT). 
     
     
         49 . The method of  claim 48 , wherein prior to the step of detecting the detectable label, the matrix is expanded. 
     
     
         50 . The methods of  claim 41 , wherein one or more of the secondary probes comprises a detectable label. 
     
     
         51 . The methods of  claim 50 , further comprising detecting the detectable label. 
     
     
         52 . The method of  claim 51 , wherein the detectable label comprises a fluorescent label, and wherein the step of detecting the detectable label comprises one or more of fluorescence microscopy, super-resolution microscopy, Stimulated Emission Depletion Microscopy (STED), structured Illumination Microscopy (SIM), and single-molecule super-resolution microscopy technologies selected from Photo-Activated Localization Microscopy (PALM), stochastic Optical Reconstruction Microscopy (STORM) and DNA-based Point Accumulation In Nanoscale Topography (DNA-PAINT). 
     
     
         53 . The method of  claim 52 , wherein prior to the step of detecting the detectable label, the matrix is expanded.