3D mapping and accelerated super-resolution imaging of the human genome using in situ sequencing

Huy Q. Nguyen, Shyamtanu Chattoraj, David Castillo, Son C. Nguyen, Guy Nir, Antonios Lioutas, Elliot A. Hershberg, Nuno M.C. Martins, Paul L. Reginato, Mohammed Hannan, Brian J. Beliveau, George M. Church, Evan R. Daugharthy, Marc A. Marti-Renom, C. ting Wu

Research output: Contribution to journalArticlepeer-review

99 Scopus citations

Abstract

There is a need for methods that can image chromosomes with genome-wide coverage, as well as greater genomic and optical resolution. We introduce OligoFISSEQ, a suite of three methods that leverage fluorescence in situ sequencing (FISSEQ) of barcoded Oligopaint probes to enable the rapid visualization of many targeted genomic regions. Applying OligoFISSEQ to human diploid fibroblast cells, we show how four rounds of sequencing are sufficient to produce 3D maps of 36 genomic targets across six chromosomes in hundreds to thousands of cells, implying a potential to image thousands of targets in only five to eight rounds of sequencing. We also use OligoFISSEQ to trace chromosomes at finer resolution, following the path of the X chromosome through 46 regions, with separate studies showing compatibility of OligoFISSEQ with immunocytochemistry. Finally, we combined OligoFISSEQ with OligoSTORM, laying the foundation for accelerated single-molecule super-resolution imaging of large swaths of, if not entire, human genomes.

Original languageEnglish
Pages (from-to)822-832
Number of pages11
JournalNature Methods
Volume17
Issue number8
DOIs
StatePublished - 1 Aug 2020
Externally publishedYes

Bibliographical note

Publisher Copyright:
© 2020, The Author(s), under exclusive licence to Springer Nature America, Inc.

Funding

We acknowledge members of the Marti-Renom and Wu laboratories for technical and conceptual support, especially T. Ryu, A. Lioutas and S. Aufmkolk as well as J. AlHaj Abed, S. D. Lee, J. Erceg and T. Hatkevich; B. Beliveau, H. Sasaki, J. Horrell, L. Cai, J. Kishi and P. Soler-Vila for discussion; D. Barclay, R. Kohman, E. Iyer, K. Rodgers, A. Skrynnyk, J. Tam and R. Terry for discussion about FISSEQ and sequencing reagents; S. Alon, F. Chen, Z. Chiang, D. Goodwin, A. Payne, A. Sinha and O. Wassie for discussion about FISSEQ; C. Ebeling, J. Rosenberg and J. Stuckey for discussion and technical assistance; F. Pan and A. Hutchinson for assistance in procuring SOLiD reagents; P. Montero-Llopis and the MicRoN imaging core at Harvard Medical School; the ImageJ discussion forum; and StackOverflow. This work was supported by a Damon Runyon Dale F. Frey Breakthrough Award (to B.J.B.) to support B.J.B. and E.A.H., awards from the NSERC of Canada (PGS D) to P.L.R., the NIH (HG005550 and HG008525) and NSF (DGE1144152) to E.R.D., the European Research Council under the Seventh Framework Program (FP7/2007–2013 609989), the European Union’s Horizon 2020 Research and Innovation Program (676556) and the Spanish Ministerio de Ciencia, Innovación y Universidades (BFU2017-85926-P) to M.A.M.-R., the Centro de Excelencia Severo Ochoa 2013–2017 (SEV-2012-0208) and the CERCA Programme/Generalitat de Catalunya to the CRG, from the NIH to GMC (RM1HG008525-03) and the NIH (DP1GM106412, R01HD091797 and R01GM123289) to C.-t.W. Micrograph images for publication figures were post-processed using brightness and contrast enhancement (ImageJ>Image>Adjust>Brightness/contrast). GraphPad Prism was also used for graphs. Molecular graphics and analyses were performed with UCSF Chimera, developed by the Resource for Biocomputing, Visualization and Informatics at the University of California and supported by the National Institutes of Health (NIH; grant no. P41-GM103311; ref. 65).

FundersFunder number
Centro de Excelencia Severo OchoaSEV-2012-0208
Damon Runyon Dale F. Frey Breakthrough Award
European Union’s Horizon 2020 research and innovation program676556
Spanish Ministerio de Ciencia, Innovación y UniversidadesBFU2017-85926-P
National Science FoundationDGE1144152
National Institutes of HealthR01GM123289, RM1HG008525-03, DP1GM106412, HG005550
National Institute of General Medical Sciences
National Institute of Child Health and Human DevelopmentR01HD091797
FP7 Ideas: European Research CouncilFP7/2007-2013 609989
Natural Sciences and Engineering Research Council of Canada
European Commission
Canadian Network for Research and Innovation in Machining Technology, Natural Sciences and Engineering Research Council of CanadaPGS D
Generalitat de Catalunya
Seventh Framework ProgrammeFP7/2007–2013 609989

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