Quantitative methods for optimizing the permeabilization of cellular tissues for spatial transcriptomics are provided. Also provided is an instrument for quantitatively optimizing the permeabilization of cellular tissues used for spatial transcriptomics.

Quantitative methods for optimizing the permeabilization of cellular tissues for spatial transcriptomics are provided. Also provided is an instrument for quantitatively optimizing the permeabilization of cellular tissues used for spatial transcriptomics.

A method of forming a polymer matrix array includes treating a surface within a well of a well array with a surface compound including a surface reactive functional group and a radical-forming distal group; applying an aqueous solution including polymer precursors to the well of the well array; and activating the radical-forming distal group of the surface coupling compound with an initiator and atom transfer radical polymerization (ATRP) catalyst to initiate radical polymerization of the polymer precursors within the well of the well array to form the polymer matrix array.

A method of forming a polymer matrix array includes treating a surface within a well of a well array with a surface compound including a surface reactive functional group and a radical-forming distal group; applying an aqueous solution including polymer precursors to the well of the well array; and activating the radical-forming distal group of the surface coupling compound with an initiator and atom transfer radical polymerization (ATRP) catalyst to initiate radical polymerization of the polymer precursors within the well of the well array to form the polymer matrix array.

Novel methods of generating a localized population of immobilized clonal amplicons on a support are provided.

Novel methods of generating a localized population of immobilized clonal amplicons on a support are provided.

An example of a kit includes a flow cell, a primer fluid, and a cleaving fluid. The flow cell includes at least one surface functionalized with a polymeric hydrogel including azide functional groups or amine functional groups. The primer fluid includes a plurality of alkyne-containing primers, each alkyne-containing primer having an amino cleavable group attaching a primer sequence of the alkyne-containing primer to an alkyne-containing moiety of the alkyne-containing primer. The cleaving fluid includes a substance that is reactive with the amino cleavable group.

An example of a kit includes a flow cell, a primer fluid, and a cleaving fluid. The flow cell includes at least one surface functionalized with a polymeric hydrogel including azide functional groups or amine functional groups. The primer fluid includes a plurality of alkyne-containing primers, each alkyne-containing primer having an amino cleavable group attaching a primer sequence of the alkyne-containing primer to an alkyne-containing moiety of the alkyne-containing primer. The cleaving fluid includes a substance that is reactive with the amino cleavable group.

In some embodiments, methods for ligating nucleic acid ends comprise: conducting a nucleic acid ligation reaction in the presence of at least one agent that generates a ligatable terminal 5′ phosphate group by removing an adenylate group from a terminal 5′ phosphate of a nucleic acid. In some embodiments, an aprataxin enzyme can catalyze removal of an adenylate group from a terminal 5′ phosphate of a nucleic acid. In some embodiments, methods for ligating nucleic acid ends comprise: conducting a nucleic acid ligation reaction in the presence of an aprataxin enzyme under conditions suitable for ligating nucleic acid ends.

In some embodiments, methods for ligating nucleic acid ends comprise: conducting a nucleic acid ligation reaction in the presence of at least one agent that generates a ligatable terminal 5′ phosphate group by removing an adenylate group from a terminal 5′ phosphate of a nucleic acid. In some embodiments, an aprataxin enzyme can catalyze removal of an adenylate group from a terminal 5′ phosphate of a nucleic acid. In some embodiments, methods for ligating nucleic acid ends comprise: conducting a nucleic acid ligation reaction in the presence of an aprataxin enzyme under conditions suitable for ligating nucleic acid ends.