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Science. 2016 Jul 29;353(6298):aaf1175. doi: 10.1126/science.aaf1175. Epub 2016 Jun 9.

Molecular recordings by directed CRISPR spacer acquisition.

Author information

1
Department of Genetics, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115, USA. Department of Stem Cell and Regenerative Biology, Center for Brain Science, and Harvard Stem Cell Institute, Harvard University, Bauer Laboratory 103, Cambridge, MA 02138, USA. Wyss Institute for Biologically Inspired Engineering, Harvard University, Cambridge, MA 02138, USA.
2
Department of Genetics, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115, USA. Wyss Institute for Biologically Inspired Engineering, Harvard University, Cambridge, MA 02138, USA.
3
Department of Stem Cell and Regenerative Biology, Center for Brain Science, and Harvard Stem Cell Institute, Harvard University, Bauer Laboratory 103, Cambridge, MA 02138, USA.
4
Department of Genetics, Harvard Medical School, 77 Avenue Louis Pasteur, Boston, MA 02115, USA. Wyss Institute for Biologically Inspired Engineering, Harvard University, Cambridge, MA 02138, USA. gchurch@genetics.med.harvard.edu.

Abstract

The ability to write a stable record of identified molecular events into a specific genomic locus would enable the examination of long cellular histories and have many applications, ranging from developmental biology to synthetic devices. We show that the type I-E CRISPR (clustered regularly interspaced short palindromic repeats)-Cas system of Escherichia coli can mediate acquisition of defined pieces of synthetic DNA. We harnessed this feature to generate records of specific DNA sequences into a population of bacterial genomes. We then applied directed evolution so as to alter the recognition of a protospacer adjacent motif by the Cas1-Cas2 complex, which enabled recording in two modes simultaneously. We used this system to reveal aspects of spacer acquisition, fundamental to the CRISPR-Cas adaptation process. These results lay the foundations of a multimodal intracellular recording device.

PMID:
27284167
PMCID:
PMC4994893
DOI:
10.1126/science.aaf1175
[Indexed for MEDLINE]
Free PMC Article

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