title: Pervasive generation of oppositely oriented spacers during CRISPR adaptation.
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.
authors.affiliation: Institute of Molecular Genetics, Russian Academy of Sciences, Moscow, 123182, Russia.
authors.forename: Sergey
authors.initials: S
authors.lastname: Shmakov
authors.affiliation: Institute of Molecular Genetics, Russian Academy of Sciences, Moscow, 123182, Russia Skolkovo Institute of Science and Technology, Skolkovo, 143025, Russia.
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Savitskaya
authors.affiliation: Waksman Institute for Microbiology, Rutgers, The State University of New Jersey, 190 Frelinghuysen Road, Piscataway, NJ 08854, USA.
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Semenova
authors.affiliation: M.V. Lomonosov Moscow State University, Moscow, 119991, Russia.
authors.forename: Maria D
authors.initials: MD
authors.lastname: Logacheva
authors.affiliation: Purdue University, West Lafayette, IN 47907, USA.
authors.forename: Kirill A
authors.initials: KA
authors.lastname: Datsenko
authors.affiliation: St Petersburg State Polytechnical University, St Petersburg, 195251, Russia severik@waksman.rutgers.edu.
authors.forename: Konstantin
authors.initials: K
authors.lastname: Severinov
mesh_terms: D001483:Base Sequence; D064112:Clustered Regularly Interspaced Short Palindromic Repeats; D017124:Conserved Sequence; D004269:DNA, Bacterial / Q000235:genetics; D021901:DNA, Intergenic; D004926:Escherichia coli / Q000235:genetics*; D019143:Evolution, Molecular
chemical_list: D004269:DNA, Bacterial; D021901:DNA, Intergenic
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