US 11,959,133 B2Granted

Methods and compositions for generating reference maps for nanopore-based polymer analysis

AssigneeUniversity of Washington through its Center for Commercialization

InventorsJens Gundlach, Ian M. Derrington, Andrew Laszlo, Elizabeth Manrao

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Abstract

The present disclosure generally relates to the methods and compositions to efficiently analyze polymer characteristics using nanopore-based assays. Specifically disclosed is a method for generating reference signals for polymer analysis in a nanopore system, wherein the nanopore system has a multi-subunit output signal resolution. The method comprises translocating a reference sequence through a nanopore to generate a plurality of reference output signals, wherein each possible multi-subunit sequence that can determine an output signal appears only once in the reference sequence. The output signals are compiled into a reference map for nanopore analysis of an analyte polymer. Also provided are methods and compositions for calibrating the nanopore system for optimized polymer analysis.

Claims12

Dependent claims

A nanopore system for sequencing an unknown polymer, the system comprising:

(a) a nanopore positioned in a nonconductive barrier between a first conductive liquid medium and a second conductive liquid medium that are in liquid communication through the nanopore;
(b) a voltage source configured to apply a voltage across the nonconductive barrier;
(c) a data acquisition device configured to detect an ion current through the nanopore;
and
(d) one or more unknown polymers operative to pass through the nanopore of the nanopore system from the first conductive liquid medium to the second conductive liquid medium to generate an ion current level pattern;
and
(e) a reference map comprising ion current level signals generated from polymer subunit combinations of a 4 or more-mer sequence;
and
wherein comparing the ion current level in the unknown polymer to the reference map provides polymer sequence information.

6 more elements

Dependencies are inferred from the claim text.

Facts

Application: 16/824,186Filed 19 Mar 2020 · continuation
Priority: 19 Apr 2012
Published: US 2020/0377936 A13 Dec 2020
Granted: 16 Apr 2024
Term ends: Not estimatedThe source records don’t support an estimate.

Classification5 codes

G01N 33/48721
By electrical means: investigating individual macromolecules, e.g. by translocation through nanopores
C12Q 1/6869
Involving nucleic acids: methods for sequencing
G01N 27/447
Systems: using electrophoresis
G01N
Investigating or analysing materials by determining their chemical or physical properties

Family

Application61/635,768Us provisional application · filed 19 Apr 2012
PCT/US2013/037454Parent of this continuation · filed 19 Apr 2013
US 2020/0377936 A1Pre-grant publication · published 3 Dec 2020
Application14/395,793Parent of this continuation
US 10,612,083 B2Parent of this continuation
US 2015/0132745 A1Published application
US 11,959,133 B2This patent

From the US filing history. Foreign family members aren’t listed.