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Geurts, Aron M.; Wilber, Andrew; Carlson, Corey M.; Lobitz, Paul D.; Clark, Karl J.; Hackett, Perry B.; McIvor, R. Scott; Largaespada, David A.

doi:10.1186/1472-6750-6-30

Cited by 38 publications

(34 citation statements)

References 45 publications

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“…The Sleeping Beauty (SB) gene-trap transposon method was used to create the trpc4 -KO animals 7 . The SB method uses cut-and-paste transposable elements to generate heritable loss-of-function mutations.…”

Section: Resultsmentioning

confidence: 99%

Cocaine self-administration in rats lacking a functional trpc4 gene

et al. 2013

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The canonical transient receptor potential (TRPC) family of Ca 2+ permeable, non-selective cation channels is abundantly expressed throughout the brain, and plays a pivotal role in modulating cellular excitability. Unlike other TRPC channels, TRPC4 subtype expression in the adult rodent brain is restricted to a network of structures that receive dopaminergic innervation, suggesting an association with motivation- and reward-related behaviors. We hypothesized that these channels may play a critical role in dopamine-dependent drug-seeking behaviors. Here, we gathered data testing trpc4 knockout (KO) rats and wild-type (WT) littermates in the acquisition of a natural sucrose reward (10 days), and cocaine self-administration (13 days) at 0.5 mg/kg/infusion. Rats lacking the trpc4 gene ( trpc4-KO) learned to lever press for sucrose to a similar degree as their WT controls. However, when they were switched to cocaine, the trpc4-KO rats had substantially reduced cocaine-paired lever pressing compared to WT controls. No obvious group differences in inactive lever pressing were observed, for any time, during cocaine self-administration.

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Section: Resultsmentioning

confidence: 99%

Cocaine self-administration in rats lacking a functional trpc4 gene

et al. 2013

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“…We show in the current study that the T2/OncZ system promotes a high level of transposition and genome-wide integration. Germline mutagenesis in the mouse starting with transgene concatemers has previously been shown for both the T2/Onc transposon [41] and a gene trap T2 vector [56]. Adopting the T2/OncZ system for germline mutagenesis might increase the efficiency of insertional mutagenesis screens compared to previous transgenic methods in zebrafish [57].…”

Section: Discussionmentioning

confidence: 99%

Somatic Mutagenesis with a Sleeping Beauty Transposon System Leads to Solid Tumor Formation in Zebrafish

et al. 2011

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Large-scale sequencing of human cancer genomes and mouse transposon-induced tumors has identified a vast number of genes mutated in different cancers. One of the outstanding challenges in this field is to determine which genes, when mutated, contribute to cellular transformation and tumor progression. To identify new and conserved genes that drive tumorigenesis we have developed a novel cancer model in a distantly related vertebrate species, the zebrafish, Danio rerio. The Sleeping Beauty (SB) T2/Onc transposon system was adapted for somatic mutagenesis in zebrafish. The carp ß-actin promoter was cloned into T2/Onc to create T2/OncZ. Two transgenic zebrafish lines that contain large concatemers of T2/OncZ were isolated by injection of linear DNA into the zebrafish embryo. The T2/OncZ transposons were mobilized throughout the zebrafish genome from the transgene array by injecting SB11 transposase RNA at the 1-cell stage. Alternatively, the T2/OncZ zebrafish were crossed to a transgenic line that constitutively expresses SB11 transposase. T2/OncZ transposon integration sites were cloned by ligation-mediated PCR and sequenced on a Genome Analyzer II. Between 700–6800 unique integration events in individual fish were mapped to the zebrafish genome. The data show that introduction of transposase by transgene expression or RNA injection results in an even distribution of transposon re-integration events across the zebrafish genome. SB11 mRNA injection resulted in neoplasms in 10% of adult fish at ∼10 months of age. T2/OncZ-induced zebrafish tumors contain many mutated genes in common with human and mouse cancer genes. These analyses validate our mutagenesis approach and provide additional support for the involvement of these genes in human cancers. The zebrafish T2/OncZ cancer model will be useful for identifying novel and conserved genetic drivers of human cancers.

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“…Trapping and discovery of low expressing genes can be further facilitated by using transcriptional transactivation systems, in which an initial, low level of the gene trap reporter signal is amplified by trans -activation of a second reporter and hence made detectable. For example, a conditionally expressed tTA (tetracycline controllable) transcriptional activator has been included in an SB-based IRES-gene trap vector to amplify the trap signal by activating transcription of a reporter in zebrafish embryos and mice [64, 70]. Gene trapping was combined with a coat color marker in order to provide a method for fast and noninvasive identification of new transposition events and homozygous rats [68].…”

Section: Transposons and Functional Genomicsmentioning

confidence: 99%

Transposons As Tools for Functional Genomics in Vertebrate Models

2017

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Genetic tools and mutagenesis strategies based on transposable elements are currently under development with a vision to link primary DNA sequence information to gene functions in vertebrate models. By virtue of their inherent capacity to insert into DNA, transposons can be developed into powerful tools for chromosomal manipulations. Transposon-based forward mutagenesis screens have numerous advantages including high throughput, easy identification of mutated alleles and providing insight into genetic networks and pathways based on phenotypes. For example, the Sleeping Beauty transposon has become highly instrumental to induce tumors in experimental animals in a tissue-specific manner with the aim of uncovering the genetic basis of diverse cancers. Here, we describe a battery of mutagenic cassettes that can be applied in conjunction with transposon vectors to mutagenize genes, and highlight versatile experimental strategies for the generation of engineered chromosomes for loss-of-function as well as gain-of-function mutagenesis for functional gene annotation in vertebrate models, including zebrafish, mice and rats.

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Untitled

Cited by 38 publications

References 45 publications

Cocaine self-administration in rats lacking a functional trpc4 gene

Cocaine self-administration in rats lacking a functional trpc4 gene

Somatic Mutagenesis with a Sleeping Beauty Transposon System Leads to Solid Tumor Formation in Zebrafish

Transposons As Tools for Functional Genomics in Vertebrate Models

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