Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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Record number 548170
Title Using CAPTURE to detect spacer acquisition in native CRISPR arrays
Author(s) McKenzie, Rebecca E.; Almendros, Cristóbal; Vink, Jochem N.A.; Brouns, Stan J.J.
Source Nature protocols (2019)14. - ISSN 1754-2189 - p. 976 - 990.
DOI https://doi.org/10.1038/s41596-018-0123-5
Department(s) Microbiology
Publication type Refereed Article in a scientific journal
Publication year 2019
Abstract

CRISPR–Cas systems are able to acquire immunological memories (spacers) from bacteriophages and plasmids in order to survive infection; however, this often occurs at low frequency within a population, which can make it difficult to detect. Here we describe CAPTURE (CRISPR adaptation PCR technique using reamplification and electrophoresis), a versatile and adaptable protocol to detect spacer-acquisition events by electrophoresis imaging with high-enough sensitivity to identify spacer acquisition in 1 in 10 5 cells. Our method harnesses two simple PCR steps, separated by automated electrophoresis and extraction of size-selected DNA amplicons, thus allowing the removal of unexpanded arrays from the sample pool and enabling 1,000-times more sensitive detection of new spacers than alternative PCR protocols. CAPTURE is a straightforward method that requires only 1 d to enable the detection of spacer acquisition in all native CRISPR systems and facilitate studies aimed both at unraveling the mechanism of spacer integration and more sensitive tracing of integration events in natural ecosystems.

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