Transposition and gene disruption in the male germline of the mouse

Dupuy, Adam J.; Fritz, Sabine; Largaespada, David A.

doi:10.1002/gene.1037

Cited by 195 publications

(173 citation statements)

References 21 publications

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“…Recent work demonstrates that the Sleeping Beauty transposase can function in transgenic mice (12)(13)(14). In these experiments, chromosomally resident transposon vectors were mobilized in transgenic mice that were ubiquitously expressing the transposase (13,14) or were expressing the transposase in the male germ line (12).…”

Section: Discussionmentioning

confidence: 99%

“…Recently a transposon system, called Sleeping Beauty, was resurrected by site-directed mutagenesis of inactive elements in salmonid genomes (10). The Sleeping Beauty transposase (SB10) is active in a wide range of vertebrate cells, including human cultured cells (10), mouse somatic tissues (11), and the mouse germ line (12)(13)(14). In this report, we investigate the potential of Sleeping Beauty to function in the one-cell mouse embryo as a potential tool for insertional mutagenesis and as an adjuvant for germ-line transgenesis in general.…”

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confidence: 99%

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Mammalian germ-line transgenesis by transposition

Dupuy

Clark

Carlson

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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193

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Transposons have been used in invertebrates for transgenesis and insertional mutagens in genetic screens. We tested a functional transposon called Sleeping Beauty in the one-cell mouse embryo. In this report, we describe experiments in which transposon vectors were injected into one-cell mouse embryos with mRNA expressing the SB10 transposase enzyme. Molecular evidence of transposition was obtained by cloning of insertion sites from multiple transgenic mice produced by SB10 mRNA͞transposon coinjection. We also demonstrate germ-line transmission and expression from transposed elements. This technique has promise as a germ-line transgenesis method in other vertebrate species and for insertional mutagenesis in the mouse.transposon ͉ insertional mutagenesis ͉ Sleeping Beauty

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Mammalian germ-line transgenesis by transposition

Dupuy

Clark

Carlson

et al. 2002

Proc. Natl. Acad. Sci. U.S.A.

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193

161

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“…Notably, differences in retrotransposition timing observed for pPolII and pHSP70-2 could be harnessed for germ-line and somatic random mutagenesis applications. A similar strategy was used successfully with a DNA transposon (Dupuy et al 2001Collier et al 2005).…”

Section: Somatic Mosaicism In Mouse Models Of L1 Retrotranspositionmentioning

confidence: 99%

L1 integration in a transgenic mouse model

et al. 2005

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To study integration of the human LINE-1 retrotransposon (L1) in vivo, we developed a transgenic mouse model of L1 retrotransposition that displays de novo somatic L1 insertions at a high frequency, occasionally several insertions per mouse. We mapped 3Ј integration sites of 51 insertions by Thermal Asymmetric Interlaced PCR (TAIL-PCR). Analysis of integration locations revealed a broad genomic distribution with a modest preference for intergenic regions. We characterized the complete structures of 33 de novo retrotransposition events. Our results highlight the large number of highly truncated L1s, as over 52% (27/51) of total integrants were <1/3 the length of a full-length element. New integrants carry all structural characteristics typical of genomic L1s, including a number with inversions, deletions, and 5Ј-end microhomologies to the target DNA sequence. Notably, at least 13% (7/51) of all insertions contain a short stretch of extra nucleotides at their 5Ј end, which we postulate result from template-jumping by the L1-encoded reverse transcriptase. We propose a unified model of L1 integration that explains all of the characteristic features of L1 retrotransposition, such as 5Ј truncations, inversions, extra nucleotide additions, and 5Ј boundary and inversion point microhomologies.[Supplemental material is available online at www.genome.org.]The long interspersed nucleotide element-1 (L1) retrotransposon enjoyed tremendous evolutionary success in colonizing eukaryotic genomes (Kazazian Jr. 2004); its roughly 500,000 copies comprise ∼17% of human DNA (Lander et al. 2001). The full-length 6-kb L1 encodes a 5Ј UTR containing an internal promoter, two proteins-ORF1, a nucleic acid-binding protein with chaperone activity (Hohjoh andSinger 1996, 1997;Martin and Bushman 2001), and ORF2, a protein with endonuclease (EN) and reverse transcriptase (RT) activities (Mathias et al. 1991;Feng et al. 1996), and a 3Ј UTR ending with a poly(A) tail (Fig. 1A). Both ORF1 and ORF2 proteins are required for retrotransposition . Once transcribed and translated, L1 RNA is copied into the genome by target-primed reverse transcription (TPRT) (Luan et al. 1993;Cost et al. 2002). During TPRT, one strand of host DNA is cleaved by the EN to expose a free 3Ј-hydroxyl, which is then used by the RT as a primer in reverse transcription, copying the L1 RNA template directly into the host genome. To complete integration, the second strand of host DNA must be cleaved, RNA removed, the newly synthesized strand copied, and breaks resolved. The mechanism by which these later steps are accomplished is only beginning to be understood, with recent biochemical data from a related non-LTR retrotransposon R2 of the silkmoth, Bombyx mori, suggesting that a second R2 protein is involved in cleavage, RNA displacement, and synthesis of the second strand (Bibillo and Eickbush 2002a; Christensen and Eickbush 2005).Following essentially random integration, endogenous L1s are thought to be lost over time due to strong negative selection leading to their uneven ...

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“…SB has proven to be particularly effective as a somatic mutagen to identify oncogenes by insertional mutagenesis (Collier et al, 2005;Dupuy et al, 2005). SB has also been used for germ line mutagenesis but has been less effective in this context (Dupuy et al, 2001;Fischer et al, 2001;Horie et al, 2001). Recently a new transposon, PiggyBac (PB) isolated from the cabbage looper moth Trichoplusia ni, has been shown to be highly efficient for germ line insertional mutagenesis in mice (Ding et al, 2005).…”

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confidence: 99%

Chromosomal mobilization and reintegration of Sleeping Beauty and PiggyBac transposons

Liang

Kong

Stalker

et al. 2009

Genesis

133

123

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Summary: The Sleeping Beauty and PiggyBac DNA transposon systems have recently been developed as tools for insertional mutagenesis. We have compared the chromosomal mobilization efficiency and insertion site preference of the two transposons mobilized from the same donor site in mouse embryonic stem (ES) cells under conditions in which there were no selective constraints on the transposons' insertion sites. Compared with Sleeping Beauty, PiggyBac exhibits higher transposition efficiencies, no evidence for local hopping and a significant bias toward reintegration in intragenic regions, which demonstrate its utility for insertional mutagenesis. Although Sleeping Beauty had no detectable genomic bias with respect to insertions in genes or intergenic regions, both Sleeping Beauty and PiggyBac transposons displayed preferential integration into actively transcribed loci. genesis 47: [404][405][406][407][408] 2009.

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Transposition and gene disruption in the male germline of the mouse

Cited by 195 publications

References 21 publications

Mammalian germ-line transgenesis by transposition

Mammalian germ-line transgenesis by transposition

L1 integration in a transgenic mouse model

Chromosomal mobilization and reintegration of Sleeping Beauty and PiggyBac transposons

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