Provided herein include a method for modifying polymerase-nucleic acid complexes, including (a) providing a plurality of surface-immobilized polymerase-nucleic acid complexes in a vessel, wherein the nucleic acid includes a primed-template nucleic acid, wherein at least a subset of the surface-immobilized polymerase-nucleic acid complexes include ternary complexes further including nucleotides; and (b) washing the surface with an aqueous solution including a diol, sulfoxide or polyol, thereby removing the nucleotides from the vessel and retaining the surface-immobilized polymerase-nucleic acid complexes in the vessel.

Provided herein include a method for modifying polymerase-nucleic acid complexes, including (a) providing a plurality of surface-immobilized polymerase-nucleic acid complexes in a vessel, wherein the nucleic acid includes a primed-template nucleic acid, wherein at least a subset of the surface-immobilized polymerase-nucleic acid complexes include ternary complexes further including nucleotides; and (b) washing the surface with an aqueous solution including a diol, sulfoxide or polyol, thereby removing the nucleotides from the vessel and retaining the surface-immobilized polymerase-nucleic acid complexes in the vessel.

This application describes methods of preparing an immobilized library of tagged RNA fragments. Also described herein are a number of methods of preparing DNA and RNA sequencing libraries from a single sample. These methods can include library preparation from single cells.

This application describes methods of preparing an immobilized library of tagged RNA fragments. Also described herein are a number of methods of preparing DNA and RNA sequencing libraries from a single sample. These methods can include library preparation from single cells.

Embodiments may include a method of analyzing a nucleic acid molecule. The method may include attaching the nucleic acid molecule to a protein. The protein may be attached to a particle with a first diameter. The method may also include applying an electric field to move a first portion of the nucleic acid molecule into an aperture. The aperture may be defined by a first electrode, an insulator, and a second electrode. The aperture may have a second diameter less than the first diameter. The method may further include contacting the first portion of the nucleic acid molecule to both the first electrode and the second electrode. The method may include applying a voltage across the first electrode and the second electrode. The current through the electrodes and the portion of the nucleic acid molecule may be measured, and a nucleotide of the nucleic acid molecule may be identified.

Embodiments may include a method of analyzing a nucleic acid molecule. The method may include attaching the nucleic acid molecule to a protein. The protein may be attached to a particle with a first diameter. The method may also include applying an electric field to move a first portion of the nucleic acid molecule into an aperture. The aperture may be defined by a first electrode, an insulator, and a second electrode. The aperture may have a second diameter less than the first diameter. The method may further include contacting the first portion of the nucleic acid molecule to both the first electrode and the second electrode. The method may include applying a voltage across the first electrode and the second electrode. The current through the electrodes and the portion of the nucleic acid molecule may be measured, and a nucleotide of the nucleic acid molecule may be identified.

Provided herein is technology relating to depositing and/or placing a macromolecule at a desired site for an assay and particularly, but not exclusively, to methods and systems for transporting a macromolecule such as a protein, a nucleic acid, or a protein:nucleic acid complex to an assay site, such as the bottom of a nanopore, a nanowell, or a zero mode waveguide.

Provided herein is technology relating to depositing and/or placing a macromolecule at a desired site for an assay and particularly, but not exclusively, to methods and systems for transporting a macromolecule such as a protein, a nucleic acid, or a protein:nucleic acid complex to an assay site, such as the bottom of a nanopore, a nanowell, or a zero mode waveguide.

Described herein are variants of alpha-hemolysin having at least one mutation, such as a mutation to a positive charge. In certain examples, the mutation is selected from V149K, E287R, H35G, T109K, P151K, K147N, E111N, M113A, or combinations thereof in the mature, wild-type alpha-hemolysin amino acid sequence. The α-hemolysin variants may also include a substitution at H144A and/or a series of glycine residues spanning residues 127 to 131 of the mature, wild-type alpha hemolysin. Also provided are nanopore assemblies including the alpha-hemolysin variants, the assembly having a decreased time-to-thread. The decreased time-to-thread, for example, increases DNA sequencing efficiency and accuracy.

Described herein are variants of alpha-hemolysin having at least one mutation, such as a mutation to a positive charge. In certain examples, the mutation is selected from V149K, E287R, H35G, T109K, P151K, K147N, E111N, M113A, or combinations thereof in the mature, wild-type alpha-hemolysin amino acid sequence. The α-hemolysin variants may also include a substitution at H144A and/or a series of glycine residues spanning residues 127 to 131 of the mature, wild-type alpha hemolysin. Also provided are nanopore assemblies including the alpha-hemolysin variants, the assembly having a decreased time-to-thread. The decreased time-to-thread, for example, increases DNA sequencing efficiency and accuracy.