DNA motifs determining the efficiency of adaptation into the Escherichia coli CRISPR array

Ido Yosef, Dror Shitrit, Moran G. Goren, David Burstein, Tal Pupko, Udi Qimron*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review


Clustered regularly interspaced short palindromic repeats (CRISPR) and their associated proteins constitute a recently identified prokaryotic defense system against invading nucleic acids. DNA segments, termed protospacers, are integrated into the CRISPR array in a process called adaptation. Here, we establish a PCR-based assay that enables evaluating the adaptation efficiency of specific spacers into the type I-E Escherichia coli CRISPR array. Using this assay, we provide direct evidence that the protospacer adjacent motif along with the first base of the protospacer (5′-AAG) partially affect the efficiency of spacer acquisition. Remarkably, we identified a unique dinucleotide, 5′-AA, positioned at the 3′ end of the spacer, that enhances efficiency of the spacer's acquisition. Insertion of this dinucleotide increased acquisition efficiency of two different spacers. DNA sequencing of newly adapted CRISPR arrays revealed that the position of the newly identified motif with respect to the 5′-AAG is important for affecting acquisition efficiency. Analysis of approximately 1 million spacers showed that this motif is overrepresented in frequently acquired spacers compared with those acquired rarely. Our results represent an example of a short nonprotospacer adjacent motif sequence that affects acquisition efficiency and suggest that other as yet unknown motifs affect acquisition efficiency in other CRISPR systems as well.

Original languageEnglish
Pages (from-to)14396-14401
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number35
StatePublished - 27 Aug 2013


  • Acquisition step
  • Defense mechanism
  • Phage-host interaction


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