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Photo of Tobias Ambjörnsson

Tobias Ambjörnsson

Senior lecturer

Photo of Tobias Ambjörnsson

Super-Resolution Genome Mapping in Silicon Nanochannels

Author

  • Jonathan Jeffet
  • Asaf Kobo
  • Tianxiang Su
  • Assaf Grunwald
  • Ori Green
  • Adam Kinos
  • Eli Eisenberg
  • Tobias Ambjörnsson
  • Fredrik Westerlund
  • Elmar Weinhold
  • Doron Shabat
  • Prashant K. Purohit
  • Yuval Ebenstein
Show all

Summary, in English

Optical genome mapping in nanochannels is a powerful genetic analysis method, complementary to deoxyribonucleic acid (DNA) sequencing. The method is based on detecting a pattern of fluorescent labels attached along individual DNA molecules. When such molecules are extended in nanochannels, the labels create a fluorescent genetic barcode that is used for mapping the DNA molecule to its genomic locus and identifying large-scale variation from the genome reference. Mapping resolution is currently limited by two main factors: the optical diffraction limit and the thermal fluctuations of DNA molecules suspended in the nanochannels. Here, we utilize single-molecule tracking and super-resolution localization in order to improve the mapping accuracy and resolving power of this genome mapping technique and achieve a 15-fold increase in resolving power compared to currently practiced methods. We took advantage of a naturally occurring genetic repeat array and labeled each repeat with custom-designed Trolox conjugated fluorophores for enhanced photostability. This model system allowed us to acquire extremely long image sequences of the equally spaced fluorescent markers along DNA molecules, enabling detailed characterization of nanoconfined DNA dynamics and quantitative comparison to the Odijk theory for confined polymer chains. We present a simple method to overcome the thermal fluctuations in the nanochannels and exploit single-step photobleaching to resolve subdiffraction spaced fluorescent markers along fluctuating DNA molecules with ∼100 bp resolution. In addition, we show how time-averaging over just ∼50 frames of 40 ms enhances mapping accuracy, improves mapping P-value scores by 3 orders of magnitude compared to nonaveraged alignment, and provides a significant advantage for analyzing structural variations between DNA molecules with similar sequence composition.

Department/s

  • Atomic Physics
  • Computational Biology and Biological Physics - Has been reorganised

Publishing year

2016-11-22

Language

English

Pages

9823-9830

Publication/Series

ACS Nano

Volume

10

Issue

11

Document type

Journal article

Publisher

The American Chemical Society (ACS)

Topic

  • Nano Technology
  • Genetics
  • Biophysics
  • Other Physics Topics

Keywords

  • confined polymers
  • DNA labeling
  • nanochannels
  • optical genome mapping
  • single-molecule
  • super-resolution

Status

Published

ISBN/ISSN/Other

  • ISSN: 1936-0851