title: A transposon-based chromosomal engineering method to survey a large cis-regulatory landscape in mice.
abstract: A large cis-regulatory landscape is a common feature of vertebrate genomes, particularly at key developmental gene loci with finely tuned expression patterns. Existing genetic tools for surveying large genomic regions of interest spanning over hundreds of kilobases are limited. Here we propose a chromosomal engineering strategy exploiting the local hopping trait of the Sleeping Beauty transposon in the mouse genome. We generated embryonic stem cells with a targeted integration of the transposon vector, carrying an enhancer-detecting lacZ reporter and loxP cassette, into the developmentally critical Pax1 gene locus, followed by efficient local transpositions, nested deletion formation and derivation of embryos by tetraploid complementation. Comparative reporter expression analysis among different insertion/deletion embryos substantially facilitated long-range cis-regulatory element mapping in the genomic neighborhood and demonstrated the potential of the transposon-based approach as a versatile tool for exploration of defined genomic intervals of functional or clinical relevance, such as disease-associated microdeletions.
authors.affiliation: Center for Advanced Science and Innovation, Osaka University, Suita, Osaka, Japan. ckokubu@mr-envi.med.osaka-u.ac.jp
authors.forename: Chikara
authors.initials: C
authors.lastname: Kokubu
authors.forename: Kyoji
authors.initials: K
authors.lastname: Horie
authors.forename: Koichiro
authors.initials: K
authors.lastname: Abe
authors.forename: Ryuji
authors.initials: R
authors.lastname: Ikeda
authors.forename: Sumi
authors.initials: S
authors.lastname: Mizuno
authors.forename: Yoshihiro
authors.initials: Y
authors.lastname: Uno
authors.forename: Sanae
authors.initials: S
authors.lastname: Ogiwara
authors.forename: Masato
authors.initials: M
authors.lastname: Ohtsuka
authors.forename: Ayako
authors.initials: A
authors.lastname: Isotani
authors.forename: Masaru
authors.initials: M
authors.lastname: Okabe
authors.forename: Kenji
authors.initials: K
authors.lastname: Imai
authors.forename: Junji
authors.initials: J
authors.lastname: Takeda
mesh_terms: D000818:Animals; D033481:Chromosomes, Mammalian / Q000235:genetics*; D004251:DNA Transposable Elements / Q000235:genetics*; D021901:DNA, Intergenic / Q000235:genetics; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism; D053595:Embryonic Stem Cells / Q000166:cytology / Q000378:metabolism; D004742:Enhancer Elements, Genetic; D005818:Genetic Engineering / Q000379:methods*; D016678:Genome / Q000235:genetics; D000090322:Homeobox Protein Nkx-2.2; D018398:Homeodomain Proteins / Q000235:genetics; D051379:Mice; D016254:Mutagenesis, Insertional; D051761:Paired Box Transcription Factors / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics*; D014157:Transcription Factors / Q000235:genetics; D029961:Zebrafish Proteins
chemical_list: D004251:DNA Transposable Elements; D021901:DNA, Intergenic; D000090322:Homeobox Protein Nkx-2.2; D018398:Homeodomain Proteins; D051761:Paired Box Transcription Factors; D014157:Transcription Factors; D029961:Zebrafish Proteins; C070835:PAX1 transcription factor