Pervasive generation of oppositely oriented spacers during CRISPR adaptation

Sergey Shmakov, Ekaterina Savitskaya, Ekaterina Semenova, Maria D. Logacheva, Kirill A. Datsenko, Konstantin Severinov

Research output: Contribution to journalArticlepeer-review

50 Scopus citations

Abstract

During the process of prokaryotic CRISPR adaptation, a copy of a segment of foreign deoxyribonucleic acid referred to as protospacer is added to the CRISPR cassette and becomes a spacer. When a protospacer contains a neighboring target interference motif, the specific small CRISPR ribonucleic acid (crRNA) transcribed from expanded CRISPR cassette can protect a prokaryotic cell from virus infection or plasmid transformation and conjugation. We show that in Escherichia coli, a vast majority of plasmid protospacers generate spacers integrated in CRISPR cassette in two opposing orientations, leading to frequent appearance of complementary spacer pairs in a population of cells that underwent CRISPR adaptation. When a protospacer contains a spacer acquisition motif AAG, spacer orientation that generates functional protective crRNA is strongly preferred. All other protospacers give rise to spacers oriented in both ways at comparable frequencies. This phenomenon increases the repertoire of available spacers and should make it more likely that a protective crRNA is formed as a result of CRISPR adaptation.

Original languageEnglish (US)
Pages (from-to)5907-5916
Number of pages10
JournalNucleic acids research
Volume42
Issue number9
DOIs
StatePublished - May 2014
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Genetics

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