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A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast

Nicholas J. McGlincy, Zuriah A. Meacham, Kendra Swain, Ryan Muller, Rachel Baum, View ORCID ProfileNicholas T. Ingolia
doi: https://doi.org/10.1101/2020.03.11.988105
Nicholas J. McGlincy
1Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
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Zuriah A. Meacham
1Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
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Kendra Swain
1Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
2Biophysics Graduate Group, University of California, Berkeley, Berkeley, CA 94720, USA
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Ryan Muller
1Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
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Rachel Baum
1Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
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Nicholas T. Ingolia
1Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA
2Biophysics Graduate Group, University of California, Berkeley, Berkeley, CA 94720, USA
3California Institute for Quantitative Biosciences, University of California, Berkeley, Berkeley, CA 94720, USA
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  • ORCID record for Nicholas T. Ingolia
  • For correspondence: ingolia@berkeley.edu
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Abstract

CRISPR/Cas9-mediated transcriptional interference (CRISPRi) enables programmable gene knock-down, yielding interpretable loss-of-function phenotypes for nearly any gene. Effective, inducible CRISPRi has been demonstrated in budding yeast, but no genome-scale guide libraries have been reported. We present a comprehensive yeast CRISPRi library, based on empirical design rules, containing 10 distinct guides for most genes. Competitive growth after pooled transformation revealed strong fitness defects for most essential genes, verifying that the library provides comprehensive genome coverage. We used the relative growth defects caused by different guides targeting essential genes to further refine yeast CRISPRi design rules. In order to obtain more accurate and robust guide abundance measurements in pooled screens, we link guides with random nucleotide barcodes and carry out linear amplification by in vitro transcription. Taken together, we demonstrate a broadly useful platform for comprehensive, high-precision CRISPRi screening in yeast.

Footnotes

  • https://ingolia-lab.github.io/yeast-crispri/

Copyright 
The copyright holder for this preprint is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity. It is made available under a CC-BY-NC 4.0 International license.
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Posted March 12, 2020.
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A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast
Nicholas J. McGlincy, Zuriah A. Meacham, Kendra Swain, Ryan Muller, Rachel Baum, Nicholas T. Ingolia
bioRxiv 2020.03.11.988105; doi: https://doi.org/10.1101/2020.03.11.988105
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A genome-scale CRISPR interference guide library enables comprehensive phenotypic profiling in yeast
Nicholas J. McGlincy, Zuriah A. Meacham, Kendra Swain, Ryan Muller, Rachel Baum, Nicholas T. Ingolia
bioRxiv 2020.03.11.988105; doi: https://doi.org/10.1101/2020.03.11.988105

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