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Nat Methods. 2017 Feb;14(2):195-200. doi: 10.1038/nmeth.4108. Epub 2016 Dec 5.

Rapidly evolving homing CRISPR barcodes.

Author information

1
Department of Genetics, Harvard Medical School, Boston, Massachusetts, USA.
2
Department of Bioengineering, University of California San Diego, La Jolla, California, USA.
3
Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, Massachusetts, USA.

Abstract

We present an approach for engineering evolving DNA barcodes in living cells. A homing guide RNA (hgRNA) scaffold directs the Cas9-hgRNA complex to the DNA locus of the hgRNA itself. We show that this homing CRISPR-Cas9 system acts as an expressed genetic barcode that diversifies its sequence and that the rate of diversification can be controlled in cultured cells. We further evaluate these barcodes in cell populations and show that they can be used to record lineage history and that the barcode RNA can be amplified in situ, a prerequisite for in situ sequencing. This integrated approach will have wide-ranging applications, such as in deep lineage tracing, cellular barcoding, molecular recording, dissecting cancer biology, and connectome mapping.

PMID:
27918539
PMCID:
PMC5322472
DOI:
10.1038/nmeth.4108
[Indexed for MEDLINE]
Free PMC Article

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