Multiplexed in vivo homology-directed repair and tumor barcoding enables parallel quantification of Kras variant oncogenicity.

Autor:	Winters IP; Department of Genetics, Stanford University School of Medicine, Stanford, CA, 94305, USA., Chiou SH; Department of Genetics, Stanford University School of Medicine, Stanford, CA, 94305, USA., Paulk NK; Department of Genetics, Stanford University School of Medicine, Stanford, CA, 94305, USA.; Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, 94305, USA., McFarland CD; Department of Biology, Stanford University, Stanford, CA, 94305, USA., Lalgudi PV; Department of Genetics, Stanford University School of Medicine, Stanford, CA, 94305, USA., Ma RK; Department of Genetics, Stanford University School of Medicine, Stanford, CA, 94305, USA., Lisowski L; Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, 94305, USA.; Translational Vectorology Group, Children's Medical Research Institute, Westmead, NSW, 2145, Australia.; Military Institute of Hygiene and Epidemiology, Puławy, 24-100, Poland., Connolly AJ; Department of Pathology, Stanford University School of Medicine, Stanford, CA, 94305, USA., Petrov DA; Department of Biology, Stanford University, Stanford, CA, 94305, USA., Kay MA; Department of Genetics, Stanford University School of Medicine, Stanford, CA, 94305, USA.; Department of Pediatrics, Stanford University School of Medicine, Stanford, CA, 94305, USA., Winslow MM; Department of Genetics, Stanford University School of Medicine, Stanford, CA, 94305, USA. mwinslow@stanford.edu.; Department of Pathology, Stanford University School of Medicine, Stanford, CA, 94305, USA. mwinslow@stanford.edu.; Cancer Biology Program, Stanford University School of Medicine, Stanford, CA, 94305, USA. mwinslow@stanford.edu.; Stanford Cancer Institute, Stanford University School of Medicine, Stanford, CA, 94305, USA. mwinslow@stanford.edu.
Jazyk:	angličtina
Zdroj:	Nature communications [Nat Commun] 2017 Dec 12; Vol. 8 (1), pp. 2053. Date of Electronic Publication: 2017 Dec 12.
DOI:	10.1038/s41467-017-01519-y
Abstrakt:	Large-scale genomic analyses of human cancers have cataloged somatic point mutations thought to initiate tumor development and sustain cancer growth. However, determining the functional significance of specific alterations remains a major bottleneck in our understanding of the genetic determinants of cancer. Here, we present a platform that integrates multiplexed AAV/Cas9-mediated homology-directed repair (HDR) with DNA barcoding and high-throughput sequencing to simultaneously investigate multiple genomic alterations in de novo cancers in mice. Using this approach, we introduce a barcoded library of non-synonymous mutations into hotspot codons 12 and 13 of Kras in adult somatic cells to initiate tumors in the lung, pancreas, and muscle. High-throughput sequencing of barcoded Kras HDR alleles from bulk lung and pancreas reveals surprising diversity in Kras variant oncogenicity. Rapid, cost-effective, and quantitative approaches to simultaneously investigate the function of precise genomic alterations in vivo will help uncover novel biological and clinically actionable insights into carcinogenesis.
Databáze:	MEDLINE
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