High-Content Analysis of CRISPR-Cas9 Gene-Edited Human Embryonic Stem Cells

Stem Cell Reports
Jared Carlson-StevermerKrishanu Saha

Abstract

CRISPR-Cas9 gene editing of human cells and tissues holds much promise to advance medicine and biology, but standard editing methods require weeks to months of reagent preparation and selection where much or all of the initial edited samples are destroyed during analysis. ArrayEdit, a simple approach utilizing surface-modified multiwell plates containing one-pot transcribed single-guide RNAs, separates thousands of edited cell populations for automated, live, high-content imaging and analysis. The approach lowers the time and cost of gene editing and produces edited human embryonic stem cells at high efficiencies. Edited genes can be expressed in both pluripotent stem cells and differentiated cells. This preclinical platform adds important capabilities to observe editing and selection in situ within complex structures generated by human cells, ultimately enabling optical and other molecular perturbations in the editing workflow that could refine the specificity and versatility of gene editing.

References

Nov 2, 2006·Genome Biology·Anne E CarpenterDavid M Sabatini
Jan 5, 2013·Science·Prashant MaliGeorge M Church
Jul 23, 2013·Nature Biotechnology·Patrick D HsuFeng Zhang
Aug 3, 2013·Nucleic Acids Research·Luhan YangGeorge Church
Oct 22, 2013·Nature Methods·Emanuel J P NazarethPeter W Zandstra
Dec 18, 2013·Science·Tim WangEric S Lander
Dec 18, 2013·Science·Ophir ShalemFeng Zhang
Feb 11, 2014·Nature Methods·Yuichiro MiyaokaBruce R Conklin
Apr 9, 2014·Journal of Biomolecular Screening·Shantanu SinghAuguste Genovesio
Jun 16, 2014·Nature Chemical Biology·Corynn KasapTarun M Kapoor
Jun 16, 2014·Nature Chemical Biology·Yegor SmurnyyYan Feng
Jun 28, 2014·Nature Methods·Sangsu BaeJin-Soo Kim
Jul 31, 2014·Nature Methods·Neville E SanjanaFeng Zhang
Nov 22, 2014·Nucleic Acids Research·Susan M ByrneGeorge M Church
Feb 3, 2015·Nature Biotechnology·Bernd ZetscheFeng Zhang
Feb 25, 2015·JAMA : the Journal of the American Medical Association·Jennifer A Doudna
Feb 26, 2015·Proceedings of the National Academy of Sciences of the United States of America·Hanhui MaThoru Pederson
Apr 8, 2015·Nature Chemical Biology·Kevin M DavisDavid R Liu
Apr 24, 2015·Journal of the American Chemical Society·James HemphillAlexander Deiters
Apr 25, 2015·Cell·Diego S D'AstolfoNiels Geijsen
May 12, 2015·Nature Biotechnology·Junwei ShiChristopher R Vakoc
May 13, 2015·Nature Biotechnology·Katrine S BosleyQi Zhou
May 23, 2015·Molecular Cell·Samuel H Sternberg, Jennifer A Doudna
May 25, 2015·Journal of Biotechnology·Xiquan LiangJonathan D Chesnut
Jun 2, 2015·Nature Methods·David M ShechnerJohn L Rinn
Jun 13, 2015·Genome Research·Han XuX Shirley Liu
Jun 16, 2015·Nature Biotechnology·Yuta NihongakiMoritoshi Sato
Jul 29, 2015·Proceedings of the National Academy of Sciences of the United States of America·Kathrin SchumannAlexander Marson
Aug 4, 2015·Stem Cell Reports·Alex LaperleKristyn S Masters

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Citations

Jun 28, 2016·International Journal of Molecular Sciences·Sylvia Merkert, Ulrich Martin
Apr 8, 2017·Expert Opinion on Drug Discovery·Quinn LuDavid J Dow
Mar 4, 2020·Science and Engineering Ethics·Qosay A E Al-BalasWael K Al-Delaimy
Aug 25, 2016·Oncotarget·Dandan YuZhenhe Suo
Sep 7, 2019·APL Bioengineering·Matthew DiSalvoNancy L Allbritton
Aug 26, 2017·Biotechnology Journal·Nicole J PiscopoKrishanu Saha
Dec 10, 2020·Nature Communications·Jared Carlson-StevermerKrishanu Saha
Jul 18, 2017·Trends in Biotechnology·Amr A Abdeen, Krishanu Saha
Nov 9, 2019·Biophysical Journal·Kaivalya MoluguKrishanu Saha

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Methods Mentioned

BETA
PCR
in vitro transcription
transfection
flow cytometry
electrophoresis
flow
transgenic
fluorescence-activated cell sorting
Hi-Seq
fluorescence microscopy

Software Mentioned

CellProfiler
MySQL
MySQL Workbench
ArrayEdit

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