Using CAPTURE to detect spacer acquisition in native CRISPR arrays

Journal article (2019)

Authors

R. McKenzie BN/Stan Brouns Lab - AS , Kavli institute of nanoscience Delft
C. Almendros Romero BN/Stan Brouns Lab - AS , Kavli institute of nanoscience Delft
J.N.A. Vink BN/Stan Brouns Lab - AS , Kavli institute of nanoscience Delft
S.J.J. Brouns BN/Stan Brouns Lab - AS , Wageningen University & Research, Kavli institute of nanoscience Delft
Research Group
BN/Stan Brouns Lab (AS) (TU Delft)
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Published Date

2019

Language

English

Reuse Rights

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Faculty

Applied Sciences

Department

BN/Bionanoscience

Research Group

BN/Stan Brouns Lab

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.