Generation of dual-gRNA library for combinatorial CRISPR screening of synthetic lethal gene pairs

Shan Tang; Xue Wu; Jinghui Liu; Qiongsi Zhang; Xinyi Wang; Shuai Shao; Birkan Gokbag; Kunjie Fan; Xiaoqi Liu; Fuhai Li; Lijun Cheng; Lang Li

doi:10.1016/j.xpro.2022.101556

Generation of dual-gRNA library for combinatorial CRISPR screening of synthetic lethal gene pairs

Shan Tang
, Xue Wu
, Jinghui Liu
, Qiongsi Zhang
, Xinyi Wang
, Shuai Shao
, Birkan Gokbag
, Kunjie Fan
, Xiaoqi Liu
, Fuhai Li
, Lijun Cheng
, Lang Li

Producción científica: Article › revisión exhaustiva

9 Citas (Scopus)

Resumen

Combinatorial CRISPR screening is useful for investigating synthetic lethality (SL) gene pairs. Here, we detail the steps for dual-gRNA library construction, with the introduction of two backbones, LentiGuide_DKO and LentiCRISPR_DKO. We describe steps for in vitro screening with 22Rv1-Cas9 and SaOS2-Cas9 cells followed by sequencing and data analysis. By introducing two backbones, we optimized the library construction process, facilitated standard pair-end sequencing, and provided options of screening on cells with or without modification of Cas9 expression.

Idioma original	English
Número de artículo	101556
Publicación	STAR Protocols
Volumen	3
N.º	3
DOI	https://doi.org/10.1016/j.xpro.2022.101556
Estado	Published - sept 16 2022

Nota bibliográfica

Publisher Copyright:
© 2022

Financiación

The work related to cell culture is done by Dr. Liu’s lab and was supported by NIH R01 CA157429 (X. Liu), R01 CA196634 (X. Liu), R01 CA264652 (X. Liu), R01 CA256893 (X. Liu).

Financiadores	Número del financiador
National Institutes of Health (NIH)	R01 CA256893, R01 CA196634, R01 CA157429
National Institutes of Health (NIH)
National Childhood Cancer Registry – National Cancer Institute	R01CA264652
National Childhood Cancer Registry – National Cancer Institute

ASJC Scopus subject areas

General Neuroscience
General Biochemistry, Genetics and Molecular Biology
General Immunology and Microbiology

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10.1016/j.xpro.2022.101556

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title = "Generation of dual-gRNA library for combinatorial CRISPR screening of synthetic lethal gene pairs",

abstract = "Combinatorial CRISPR screening is useful for investigating synthetic lethality (SL) gene pairs. Here, we detail the steps for dual-gRNA library construction, with the introduction of two backbones, LentiGuide\_DKO and LentiCRISPR\_DKO. We describe steps for in vitro screening with 22Rv1-Cas9 and SaOS2-Cas9 cells followed by sequencing and data analysis. By introducing two backbones, we optimized the library construction process, facilitated standard pair-end sequencing, and provided options of screening on cells with or without modification of Cas9 expression.",

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AU - Tang, Shan

AU - Wu, Xue

AU - Liu, Jinghui

AU - Zhang, Qiongsi

AU - Wang, Xinyi

AU - Shao, Shuai

AU - Gokbag, Birkan

AU - Fan, Kunjie

AU - Liu, Xiaoqi

AU - Li, Fuhai

AU - Cheng, Lijun

AU - Li, Lang

PY - 2022/9/16

Y1 - 2022/9/16

N2 - Combinatorial CRISPR screening is useful for investigating synthetic lethality (SL) gene pairs. Here, we detail the steps for dual-gRNA library construction, with the introduction of two backbones, LentiGuide_DKO and LentiCRISPR_DKO. We describe steps for in vitro screening with 22Rv1-Cas9 and SaOS2-Cas9 cells followed by sequencing and data analysis. By introducing two backbones, we optimized the library construction process, facilitated standard pair-end sequencing, and provided options of screening on cells with or without modification of Cas9 expression.

AB - Combinatorial CRISPR screening is useful for investigating synthetic lethality (SL) gene pairs. Here, we detail the steps for dual-gRNA library construction, with the introduction of two backbones, LentiGuide_DKO and LentiCRISPR_DKO. We describe steps for in vitro screening with 22Rv1-Cas9 and SaOS2-Cas9 cells followed by sequencing and data analysis. By introducing two backbones, we optimized the library construction process, facilitated standard pair-end sequencing, and provided options of screening on cells with or without modification of Cas9 expression.

KW - Biotechnology and bioengineering

KW - CRISPR

KW - High Throughput Screening

KW - Molecular Biology

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Generation of dual-gRNA library for combinatorial CRISPR screening of synthetic lethal gene pairs

Resumen

Nota bibliográfica

Financiación

ASJC Scopus subject areas

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