TY - JOUR
T1 - Single-cell analysis of a mutant library generated using CRISPR-guided deaminase in human melanoma cells
AU - Jun, Soyeong
AU - Lim, Hyeonseob
AU - Chun, Honggu
AU - Lee, Ji Hyun
AU - Bang, Duhee
N1 - Funding Information:
This work was supported by: (i) the Mid-career Researcher Program (NRF-2018R1A2A1A05079172), (ii) the Bio & Medical Technology Development Program (NRF-2016M3A9B6948494), (iii) the Bio & Medical Technology Development Program (NRF-2018M3A9H3024850), (iv) the Bio & Medical Technology Development Program (NRF-2018M3A9D7079485), (v) Basic Science Research Program (NRF-2018R1A2B2001322) of the National Research Foundation of Korea, funded by the Ministry of Science, ICT & Planning, and (vi) Korea Health Technology R&D Project (HI18C2282) through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare.
Publisher Copyright:
© 2020, The Author(s).
PY - 2020/12/1
Y1 - 2020/12/1
N2 - CRISPR-based screening methods using single-cell RNA sequencing (scRNA-seq) technology enable comprehensive profiling of gene perturbations from knock-out mutations. However, evaluating substitution mutations using scRNA-seq is currently limited. We combined CRISPR RNA-guided deaminase and scRNA-seq technology to develop a platform for introducing mutations in multiple genes and assessing the mutation-associated signatures. Using this platform, we generated a library consisting of 420 sgRNAs, performed sgRNA tracking analysis, and assessed the effect size of the response to vemurafenib in the human melanoma cell line, which has been well-studied via knockout-based drop-out screens. However, a substitution mutation library screen has not been applied and transcriptional information for mechanisms of action was not assessed. Our platform permits discrimination of several candidate mutations that function differently from other mutations by integrating sgRNA candidates and gene expression readout. We anticipate that our platform will enable high-throughput analyses of the mechanisms related to a variety of biological events.
AB - CRISPR-based screening methods using single-cell RNA sequencing (scRNA-seq) technology enable comprehensive profiling of gene perturbations from knock-out mutations. However, evaluating substitution mutations using scRNA-seq is currently limited. We combined CRISPR RNA-guided deaminase and scRNA-seq technology to develop a platform for introducing mutations in multiple genes and assessing the mutation-associated signatures. Using this platform, we generated a library consisting of 420 sgRNAs, performed sgRNA tracking analysis, and assessed the effect size of the response to vemurafenib in the human melanoma cell line, which has been well-studied via knockout-based drop-out screens. However, a substitution mutation library screen has not been applied and transcriptional information for mechanisms of action was not assessed. Our platform permits discrimination of several candidate mutations that function differently from other mutations by integrating sgRNA candidates and gene expression readout. We anticipate that our platform will enable high-throughput analyses of the mechanisms related to a variety of biological events.
UR - http://www.scopus.com/inward/record.url?scp=85082904028&partnerID=8YFLogxK
U2 - 10.1038/s42003-020-0888-2
DO - 10.1038/s42003-020-0888-2
M3 - Article
C2 - 32242071
AN - SCOPUS:85082904028
SN - 2399-3642
VL - 3
JO - Communications Biology
JF - Communications Biology
IS - 1
M1 - 154
ER -