Bionano Genomics, Inc.

Drexel University

Temple University

The Wistar Institute

Ming H Xiao

Han Cao

Parikshit A Deshpande

Michael Boyce-Jacino

Jennifer McCaffrey

Kandaswamy Vijayan

Michael David Austin

Wenhui Hu

Alex R Hastie

Alexey Y Sharonov

Michael G Saghbini

Henry B Sadowski

Justin Sibert

Harold C Riethman

Alexy Y Sharonov

Methods for single-molecule preparation and analysis are disclosed herein. The methods can, for example, be used for isolating and analyzing DNA from various biological samples.

Methods for single-molecule analysis

Provided are methods and devices for single-molecule genomic analysis. In one embodiment, the methods entail processing a double-stranded nucleic acid and characterizing said nucleic acid. These methods are useful in, e.g. determining structural variations and copy number variations between individuals.

Methods and devices for single-molecule whole genome analysis

A method of detecting the length of an individual telomere is provided. In one embodiment, the method includes contacting genomic DNA with a guide RNA having a portion complementary to a telomere repeat sequence in the genomic DNA and with Cas9 nickase to produce a single-strand break in the genomic DNA at the telomere repeat sequence. The nicked DNA is contacted with a polymerase and fluorescently labeled nucleotide, wherein the fluorescently labeled nucleotide is incorporated into the nicked DNA at the telomere repeat sequence. The genomic DNA is contacted with a second nicking endonuclease which is specific for a sequence motif in the genomic DNA thereby producing a second nick in the genomic DNA at the motif sequence. The nicked DNA is contacted with a polymerase and second fluorescently labeled nucleotide of different color, wherein the second fluorescently labeled nucleotide is incorporated into the nicked DNA at the motif sequence location. The length of the telomere is detected by measuring the fluorescence of first fluorescently labeled nucleotide at the telomere repeat location, wherein the fluorescently labeled motif sequences are used as a barcode to identify the chromosome.

Methods of specifically labeling nucleic acids using CRISPR/Cas

Provided herein are methods of detecting a target nucleic acid sequence. In one embodiment, the method includes contacting genomic DNA with a guide RNA having a portion complementary to the target sequence in the genomic DNA and with Cas9 nickase to produce a single-strand break in the genomic DNA at a specific location adjacent to the target sequence. The method further includes contacting the nicked DNA with a polymerase and fluorescently labeled nucleotide. The fluorescently labeled nucleotide is incorporated into the nicked DNA at the specific location and the target nucleic acid sequence is detected via fluorescent label.

The present invention provides methods of obtaining structural information about a biopolymer sample. The methods include labeling portions of a biopolymer, such as DNA or RNA, linearizing the biopolymer in some cases, and determining the distance between the labels. The user can then compare different samples' between-label distances to qualitatively compare different samples and to assay a given sample for additions or deletions of nucleotides in the regions flanked by the labels. The methods also permit sequencing of biopolymers.

Polynucleotide mapping and sequencing

The present invention relates to innovative means of generating sequence-linked DNA fragments and subsequent uses of such linked DNA fragments for de novo haplotype-resolved whole genome mapping and massively parallel sequencing. In various embodiments described herein, the methods of the invention relate to methods of generating paired-end nucleic acid fragment sharing common linker nucleic acid sequences using a nicking endonuclease, a T7 endonuclease, a restriction enzyme, or a transposase, methods of analyzing the nucleotides sequences from the linked-paired-end sequenced fragments, and methods of de novo whole genome mapping. Thus, the methods of this invention allow establishing sequence contiguity across the whole genome, and achieving high-quality, low-cost de novo assembly of complex genomes.

Whole genome mapping by DNA sequencing with linked-paired-end library

Provided are methods and devices for single-molecule genomic analysis. In one embodiment, the methods entail processing a double-stranded nucleic acid and characterizing said nucleic acid. These methods are useful in, e.g., determining structural variations and copy number variations between individuals.

Growing community of inventors

Huntingdon Valley, PA, United States of America

Ming H Xiao