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Proc Natl Acad Sci U S A. 2017 Jan 24;114(4):722-727. doi: 10.1073/pnas.1615735114. Epub 2017 Jan 6.

piggyBac mediates efficient in vivo CRISPR library screening for tumorigenesis in mice.

Xu C1, Qi X1, Du X1, Zou H1, Gao F1, Feng T1, Lu H1, Li S2,3, An X1, Zhang L1, Wu Y4, Liu Y2,3,5, Li N1, Capecchi MR6, Wu S7.

Author information

1
State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, China.
2
Department of Neurosurgery, Medical School, University of Texas Health Science Center at Houston, Houston, TX 77030.
3
Center for Stem Cell and Regenerative Medicine, University of Texas Health Science Center at Houston, Houston, TX 77030.
4
Department of Human Genetics, University of Utah School of Medicine, Salt Lake City, UT 84112.
5
The Senator Lloyd & B. A. Bentsen Center for Stroke Research, the Brown Foundation Institute of Molecular Medicine for the Prevention of Human Diseases, University of Texas Health Science Center at Houston, Houston, TX 77030.
6
Department of Human Genetics, University of Utah School of Medicine, Salt Lake City, UT 84112; swu@cau.edu.cn capecchi@genetics.utah.edu.
7
State Key Laboratory of Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193, China; swu@cau.edu.cn capecchi@genetics.utah.edu.

Abstract

CRISPR/Cas9 is becoming an increasingly important tool to functionally annotate genomes. However, because genome-wide CRISPR libraries are mostly constructed in lentiviral vectors, in vivo applications are severely limited as a result of difficulties in delivery. Here, we examined the piggyBac (PB) transposon as an alternative vehicle to deliver a guide RNA (gRNA) library for in vivo screening. Although tumor induction has previously been achieved in mice by targeting cancer genes with the CRISPR/Cas9 system, in vivo genome-scale screening has not been reported. With our PB-CRISPR libraries, we conducted an in vivo genome-wide screen in mice and identified genes mediating liver tumorigenesis, including known and unknown tumor suppressor genes (TSGs). Our results demonstrate that PB can be a simple and nonviral choice for efficient in vivo delivery of CRISPR libraries.

KEYWORDS:

CRISPR/Cas9; liver cancer; piggyBac transposon; screening; tumorigenesis

PMID:
28062688
PMCID:
PMC5278437
DOI:
10.1073/pnas.1615735114
[Indexed for MEDLINE]
Free PMC Article

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