Back to results
US 10,851,411 B2Granted
Molecular identification with subnanometer localization accuracy
AssigneesLudwig-Maximilians-Universitat Munchen, XGenomes Corp.
InventorsJohannes B. Wöhrstein, Kalim Mir, Ralf Jungmann, Florian Schüder
Life of this patent
Estimating the term…
Abstract
The present invention relates to methods of determining the sequence of nucleotides in target nucleic acid molecules. Thus, the invention relates to methods of sub-unit sequencing. The methods comprise the use of identification nucleic acid detection entities which specifically hybridize to the target nucleic acid, bind identification tags and have localization tags transiently bind thereto.
Claims20
A method of determining the sequence of nucleotides in a target nucleic acid molecule comprising the steps of:
- (1) providing a target nucleic acid molecule, wherein copies of the target nucleic acid molecule are immobilized on a solid substrate,
- (2) providing a plurality of nucleic acid detection entities, wherein each nucleic acid detection entity is at least in part single stranded and comprises:
- (i) a specific probe nucleotide sequence,
- (ii) a localization nucleotide sequence for transient binding of a localization tag,
- and
- (iii) an identification nucleotide sequence for stable hybridization with an identification tag specific for the specific probe nucleotide sequence (i),
- (3) providing a plurality of identification tags,
- wherein each identification tag
- is specific for a specific probe nucleotide sequence (i) of the nucleic acid detection entity, and
- comprises a nucleotide sequence complementary to the identification nucleotide sequence (iii) of the nucleic acid detection entity,
- (4) providing a plurality of localization tags,
- wherein said localization tag comprises
- a nucleotide sequence complementary to the localization nucleotide sequence (ii) of the nucleic acid detection entity, and
- marker(s) or label(s),
- (5) hybridizing and optionally ligating the nucleic acid detection entities to a single strand of the target nucleic acid molecules,
- (6) hybridizing the identification tags to the identification nucleotide sequence (iii) of the nucleic acid detection entities,
- optionally, stretching and/or aligning the identification nucleotide sequence (iii) of the nucleic acid detection entities,
- (7) detecting the identification tags,
- (8) transiently hybridizing the localization tags, to the localization nucleotide sequence (ii) of the nucleic acid detection entities and detecting said transient hybridization, and
- (9) spatially detecting and identifying the nucleic acid sequence in the target nucleic acid molecules.
17 more elements
Dependencies are inferred from the claim text.