title: Violation of the 12/23 rule of genomic V(D)J recombination is common in lymphocytes.
abstract: V(D)J genomic recombination joins single gene segments to encode an extensive repertoire of antigen receptor specificities in T and B lymphocytes. This process initiates with double-stranded breaks adjacent to conserved recombination signal sequences that contain either 12- or 23-nucleotide spacer regions. Only recombination between signal sequences with unequal spacers results in productive coding genes, a phenomenon known as the "12/23 rule." Here we present two novel genomic tools that allow the capture and analysis of immune locus rearrangements from whole thymic and splenic tissues using second-generation sequencing. Further, we provide strong evidence that the 12/23 rule of genomic recombination is frequently violated under physiological conditions, resulting in unanticipated hybrid recombinations in ∼10% of Tcra excision circles. Hence, we demonstrate that strict adherence to the 12/23 rule is intrinsic neither to recombination signal sequences nor to the catalytic process of recombination and propose that nonclassical excision circles are liberated during the formation of antigen receptor diversity.
authors.affiliation: Systems Biology Laboratory UK, Abingdon, Oxfordshire OX14 4SA, United Kingdom;
authors.forename: Nicholas J
authors.initials: NJ
authors.lastname: Parkinson
authors.affiliation: Systems Biology Laboratory UK, Abingdon, Oxfordshire OX14 4SA, United Kingdom;
authors.forename: Matthew
authors.initials: M
authors.lastname: Roddis
authors.affiliation: Systems Biology Laboratory UK, Abingdon, Oxfordshire OX14 4SA, United Kingdom;
authors.forename: Ben
authors.initials: B
authors.lastname: Ferneyhough
authors.affiliation: Systems Biology Laboratory UK, Abingdon, Oxfordshire OX14 4SA, United Kingdom;
authors.forename: Gang
authors.initials: G
authors.lastname: Zhang
authors.affiliation: Systems Biology Laboratory UK, Abingdon, Oxfordshire OX14 4SA, United Kingdom;
authors.forename: Adam J
authors.initials: AJ
authors.lastname: Marsden
authors.affiliation: Systems Biology Laboratory UK, Abingdon, Oxfordshire OX14 4SA, United Kingdom; MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT, United Kingdom;
authors.forename: Siarhei
authors.initials: S
authors.lastname: Maslau
authors.affiliation: Systems Biology Laboratory UK, Abingdon, Oxfordshire OX14 4SA, United Kingdom;
authors.forename: Yasmin
authors.initials: Y
authors.lastname: Sanchez-Pearson
authors.affiliation: Paediatric Immunology, Department of Biomedicine, University of Basel and The Basel University Children's Hospital, 4058 Basel, Switzerland;
authors.forename: Thomas
authors.initials: T
authors.lastname: Barthlott
authors.affiliation: Institute of Infection and Immunity, Cardiff University School of Medicine, Heath Park, Cardiff CF14 4XN, United Kingdom;
authors.forename: Ian R
authors.initials: IR
authors.lastname: Humphreys
authors.affiliation: Institute of Infection and Immunity, Cardiff University School of Medicine, Heath Park, Cardiff CF14 4XN, United Kingdom;
authors.forename: Kristin
authors.initials: K
authors.lastname: Ladell
authors.affiliation: Institute of Infection and Immunity, Cardiff University School of Medicine, Heath Park, Cardiff CF14 4XN, United Kingdom; Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892, USA;
authors.forename: David A
authors.initials: DA
authors.lastname: Price
authors.affiliation: MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT, United Kingdom;
authors.forename: Chris P
authors.initials: CP
authors.lastname: Ponting
authors.affiliation: Paediatric Immunology, Department of Biomedicine, University of Basel and The Basel University Children's Hospital, 4058 Basel, Switzerland; Developmental Immunology, Weatherall Institute of Molecular Medicine and Department of Paediatrics, University of Oxford, Oxford OX3 9DS, United Kingdom;
authors.forename: Georg
authors.initials: G
authors.lastname: Hollander
authors.affiliation: Systems Biology Laboratory UK, Abingdon, Oxfordshire OX14 4SA, United Kingdom; Department of Oncology, Division of Cellular and Molecular Medicine, St. George's, University of London, London SW17 0QT, United Kingdom.
authors.forename: Michael D
authors.initials: MD
authors.lastname: Fischer
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000235:genetics; D015321:Gene Rearrangement; D016654:Genes, RAG-1; D023281:Genomics; D006801:Humans; D008214:Lymphocytes / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D018345:Mice, Knockout; D008954:Models, Biological; D017422:Sequence Analysis, DNA; D060152:V(D)J Recombination