Methods and related devices for single molecule whole genome analysis

A base, part of the technology, applied in the field of nanotechnology and single-molecule genome analysis, can solve problems such as isolation, incompatibility, and cumbersome

Inactive Publication Date: 2014-01-08
BIONANO GENOMICS
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, techniques for examining each of the aforementioned aspects of a cell's molecular state are often isolated, ...

Method used

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  • Methods and related devices for single molecule whole genome analysis
  • Methods and related devices for single molecule whole genome analysis
  • Methods and related devices for single molecule whole genome analysis

Examples

Experimental program
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Embodiment

[0139] Example: Generation of Single-Stranded DNA Flaps on Double-Stranded DNA Molecules

[0140] Genomic DNA samples were diluted to 50 ng for nicking reactions. Add 10 μL λDNA (50ng / μL) to a 0.2mL PCR centrifuge tube, then add 2 μL 10X NE Buffer #2 and 3 μL nicking endonucleases, including but not limited to Nb.BbvCI, Nb.BsmI, Nb.BsrDI, Nb. BtsI, Nt.AlwI, Nt.BbvCI, Nt.BspQI, Nt.BstNBI, Nt.CviPII. The mixture was incubated at 37°C for 1 hour.

[0141] After the nicking reaction was complete, the experiment proceeded to limited polymerase extension at the nicking site to displace the 3' downstream strand and form a single-stranded flap. The flap generation reaction mix consisted of 15 μl of nicking product and 5 μl of incorporation mix containing 2 μl of 10X buffer, 0.5 μl of polymerase, and 1 μl of nucleotides at various concentrations from 1 μM to 1 mM. Enzymes include, but are not limited to, vent (exon-), Bst, and Phi29 polymerases. The flap generation reaction mixture...

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Abstract

Provided are methods of labeling and analyzing features along at least one macromolecule such as a linear biopolymer, including methods of mapping the distribution and frequency of specific sequence motifs or the chemical or proteomic modification state of such sequence motifs along individual unfolded nucleic acid molecules. The present invention also provides methods of identifying signature patterns of sequence or epigenetic variations along such labeled macromolecules for direct massive parallel single molecule level analysis. The present invention also provides systems suitable for high throughput analysis of such labeled macromolecules.

Description

[0001] Cross references to related applications [0002] This application claims priority to U.S. Application Serial No. 61 / 253,639, "Methods and Devices for Single Molecule Whole Genome Analysis," filed October 21, 2009, which The entire content of is hereby incorporated by reference. technical field [0003] The invention relates to the fields of nanotechnology and single-molecule genome analysis. Background technique [0004] Macromolecules such as DNA or RNA are long polymer chains composed of nucleotides, the linear sequence of which is directly related to the genomic and post-genomic gene expression information of the source organism. [0005] Sequence regions, motifs and functional units such as open reading frames (ORFs), untranslated regions (UTRs), exons, introns, protein factor binding sites, epigenomic sites such as CpG clusters, microRNA sites, Direct sequencing and mapping of transposons, retrotransposons, and other structural and functional units is importan...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
CPCC12Q1/683G01N21/6486C12Q2537/137C12Q2561/109C12Q2563/107C12Q2563/185C12Q2535/131C12Q1/34
Inventor 肖明索梅斯库玛·达斯
Owner BIONANO GENOMICS
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