I report here at (1) phiC31 integrase is functional in zebrafish cells, (2) phiC31 integrase can excise a transgene cassette flanked by an attB and an attP site, analogous to a common use of e Cre/lox SSR system, (3) phiC31 integrase functions in e zebrafish germline, and (4) a phiC31 integrase‐estrogen receptor hormone‐binding domain Cited by: 28. Transgene excision in zebrafish using e phiC31 integrase. James. Lister. Corresponding Au. E-mail address: [email protected] Department of Human and Molecular Genetics, Virginia Commonweal University School of Medicine, Richmond, Virginia.Cited by: 28. 01, · e PhiC31 integrase is functional in zebrafish cells and at present offers an alternative, or complement, to existing tools such as Cre and Flp for regulated and Cited by: . PhiC31 integrase induces intramolecular recombination between integrated target sites in zebrafish. a Schematic of e gene/enhancer trap vector Tg(UAS:[mCherry-T-mCitrine]. attP-hsp70 l:Gal4VP16. Feb 01, 20 · Transgene excision in zebrafish using e phiC31 integrase Transgene excision in zebrafish using e phiC31 integrase Lister, James. 20 -02-01 00:00:00 Genome engineering strategies employing site‐specific recombinases (SSRs) have become invaluable to e study of gene function in model organisms. One such SSR, e integrase encoded by e Streptomyces bacteriophage phiC31. 01, · Mutational derivatives of PhiC31 integrase wi increased efficiency and specificity. Mol er 17,112–120 [PMC free article] Lu J., Maddison L.A., and Chen W. (20). PhiC31 integrase induces efficient site-specific excision in zebrafish. Transgenic Res 20,183–189 [PMC free article]. Request PDF. Transgene excision in zebrafish using e phiC31 integrase. Single optical section of a 72 hour post-fertilization T2K-XpGbR (EF1-alpha-attP-GFP-attB-DsRed-Express) transgenic. To assay Int-phiC31 integrase activity on episomal DNA in zebrafish embryos, about 1 nl sterile isotonic saline solution containing a mixture of Tg(eab2:attP-mCitrine-attB-mCherry)DNA (30 ng/µl) wi or wi out syn etic phiC31 RNA (30 ng/µl) was injected into fertilized eggs of wild type zebrafish. ZEBRAFISH meeting attendees will have e opportunity to present eir research rough oral and poster presentations and to hear e latest updates. ZEBRAFISH organizers are confident at we will offer a strong scientific programme, including many opportunities to learn, share, network and interact wi our sponsors at e virtual exhibition. Feb 01, 20 · I report here at (1) phiC31 integrase is functional in zebrafish cells, (2) phiC31 integrase can excise a transgene cassette flanked by an attB and an attP site, analogous to a common use of e Cre/lox SSR system, (3) phiC31 integrase functions in e zebrafish germline, and (4) a phiC31 integrase-estrogen receptor hormone-binding domain. Here we show at phiC31 integrase efficiently induces site-specific deletion of episomal targets as well as chromosomal targets in zebrafish embryos. us, e phiC31 system can be used in zebrafish for genetic manipulations, expanding e repertoire of available tools for genetic manipulation in is vertebrate model. Site-directed zebrafish transgenesis into single landing sites wi e phiC31 integrase system. Mosimann C, Puller AC, Lawson KL, Tschopp P, Amsterdam A, Zon LI Dev Dyn. .242(8):949-63. doi: . 02/dvdy.23989. Epub 3. PubMed Article. Feb 01, 20 · Single optical section of a 72 hour post‐fertilization T2K‐XpGbR (EF1‐alpha‐attP‐GFP‐attB‐DsRed‐Express) transgenic zebrafish larva obtained by laser scanning confocal microscopy. Injection of phiC31 integrase messenger RNA at e one‐cell stage induces recombination of e transgene in a mosaic fashion, resulting in excision of e green fluorescent . Conclusions: Our results establish phiC31 integrase‐controlled site‐directed transgenesis into single, genomic attP sites as space‐, time‐, and labor‐efficient zebrafish transgenesis technique. e described reagents are available for distribution to e zebrafish community. In zebrafish, phiC31 integrase has been shown to catalyze efficient intramolecular DNA recombination of episomal and chromosomal targets (Lister, 20, . Lu et al., ). Zebrafish (Danio rerio) is an ideal in vivo model to study a wide variety of human cancer types. In is review, we provide a comprehensive overview of zebrafish in e cancer drug discovery process, from (i) approaches to induce malignant tumors, (ii) techniques to monitor cancer progression, and (iii) strategies for compound administration to (iv) a compilation of e 355 existing case. In zebrafish, enzymatic activity of PhiC31 integrase in somatic and germline cells has been demonstrated (Hu et al., . Lister, 20 . Lu et al., ). Recently, a site-specific transgenesis system wi characterized landing sites has been reported for zebrafish (Mosimann et al., ). Here, we present a PhiC31 integrase-based system. ), we ided to develop a PhiC31-based me od for analysis of enhancer activity in zebrafish. e natural function of PhiC31 integrase is to facilitate e integration of e phage genome into e bacterial host genome (Kuhstoss and Rao, 1991). PhiC31 integrase binds to two recognition sites, attP and attB, and exchanges DNA. 01, · In a number of species, cells contain a specific integrase, PhiC31, capable of recombining e DNA sequences attB and attP added to vectors wi similar genomic sequences wi a good efficiency. is recombination integrates e vector, and it generates attL and attR sites which are not recognized by e PhiC31 integrase. e integration of e. Site‐specific recombinases (SSRs) are powerful tools for genome manipulation, used in diverse organisms including Drosophila melanogaster, mouse, Arabidopsis, zebrafish, and human cultured cells. e integrase from e bacteriophage ΦC31 belongs to e large serine family of integrases, and in contrast to o er widely used SSRs such as Cre and Flp, recombination is directional and erefore. PhiC31 integrase is a DNA recombinase derived from Streptomyces phage φC31. is enzyme can mediate recombination between two sequences, namely, attB and attP 2,3.PhiC31 integrase does not. Site-directed zebrafish transgenesis into single landing sites wi e phiC31 integrase system Christian Mosimann1,2,3,6, Ann-Christin Puller1,2,3, Katy L. Lawson1. In zebrafish, phiC31 integrase has been shown to catalyze efficient intramolecular DNA recombination of episomal and chromosomal targets (Lister, 20 (Lister, Lu et al., ). e integrase from phiC31 has been deployed in multiple eukaryotes, including fruit flies (Gro et al. 2004), zebrafish (Mosimann et al. ), tobacco (Lutz et al. 2004), and cultured human. Feb 01, · In vivo detection system for targeted integration of reporter constructs in zebrafish. We have adapted a site-specific integration system wi PhiC31 integrase consisting of two components: ‘recipient transgenic lines’ containing docking attP site(s) in e genome, and targeting plasmids containing attB site(s) (orpe and Smi, 1998).In mouse and human cells, integrations were . 01, · Genetic analysis is facilitated by e efficient production of transgenic strains expressing a DNA of interest as a single copy at a designated chromosomal location. However, technical progress tod is goal in medaka fish (Oryzias latipes), a vertebrate model organism, has been slow. It is well known at phiC31 integrase enables efficient site-directed transgenesis by catalyzing e. Skip to Article Content. Skip to Article Information. Featured community. Creative Data Solutions (CDS) is a Vanderbilt Shared Resource and has extensive experience in providing effective and robust solutions to challenges pertaining to research data using modern informatics and bioinformatics approaches. AREAS COVERED: e phiC31 integrase system has been under development for ten years and has been demonstrated to be effective for integration of plasmids in a variety of tissues and organs for gene erapy in animal systems, as well as in isolated human cells. We focus on work wi e phiC31 integrase system during e past 12 - 18 mon s. e potential use of e PhiC31 integrase system in gene erapy opens up e possibilities of new treatments for old diseases. PhiC31 integrase mediates e integration of plasmid DNA into e chromsomes of mammalian cells in a sequence-specific manner, resulting in . Plasmid pcDNA3.1 phiC31 from Dr. Konrad Basler's lab contains e insert phiC31 and is published in Proc Natl Acad Sci U S. 2007 Feb 27. 4(9):3312-7. Epub 2007 Feb 22. is plasmid is available rough Addgene. Site specific recombinases (SSRs) are powerful tools for genome manipulation, used in diverse organisms including Drosophila melanogaster, mouse, Arabidopsis, zebrafish, and human cultured cells. e integrase from e bacteriophage ΦC31 belongs to e large serine family of integrases, and in contrast to o er widely used SSRs such as Cre and Flp, recombination is directional and erefore. 30, · Over e past ade, e integrase enzyme from phage phiC31 has proven to be a useful genome engineering tool in a wide variety of species, including mammalian cells. e enzyme efficiently mediates recombination between two distinct sequences, attP and attB, producing recombinant product sites, attL and attR. Hu G, Goll MG, Fisher S () PhiC31 integrase mediates efficient cassette exchange in e zebrafish germline. Dev Dyn 240(9):2 1–2 7. doi: . 02/dvdy.22699 CrossRef PubMed PubMedCentral Google . Fig. 1 In vivo detection system for targeted integration of reporter constructs in zebrafish. (A) Schematic of PhiC31 targeted integration system. Transgenic embryos containing an attP docking site have a green lens due to e gamma-crystalline promoter driving a GFP reporter. ITR labels recognition sequences of ei er Tol2 or Sleeping beauty transposases used in generating e recipient. aimed to establish a PhiC31 integrase system of targeted transgenesis in zebrafish. Firstly, we designed and tested a multicomponent system including a recipient vector, donor vector and PhiC31integrase mRNA. ese components were co-injected into one cell stage embryos to test e integration function of PhiC31 integrase in zebrafish. e. Site-directed zebrafish transgenesis into single landing sites wi e phiC31 integrase system. Mosimann C, Puller AC, Lawson KL, Tschopp P, Amsterdam A, Zon LI. Dev Dyn. .242(8):949-63. doi: . 02/dvdy.23989. In zebrafish, enzymatic activity of PhiC31 integrase in somatic and germline cells has been demonstrated (Hu et al., . Lister, 20 . Lu et al., ). Recently, a site-specific transgenesis system wi characterized landing sites has been reported for zebrafish (Mosimann et al., ). Here, we present a PhiC31 integrase-based. e phiC31 integrase enzyme made it possible to add genes, but it was not possible to remove e genes, if desired. In is new study, we show at a newly-identified protein, called excisionase or recombination directionality factor, can reverse e recombination reaction at is carried out by phiC31 integrase. Citation: Kapusi E, Kempe K, Rubtsova M, Kumlehn J, Gils M (phiC31 Integrase-Mediated Site-Specific Recombination in Barley Eszter Kapusi. 0 Katja Kempe. 0 Myroslava Rubtsova 0 Jochen Kumlehn 0 io Gils 0 Leandro Pe na, Instituto Valenciano De Investigaciones Agrarias, Spain 0 Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben, Gatersleben, Germany e. 16, · Site-specific integration has been widely used in a variety of species, including bacterium, fly, zebrafish and mammals [15,16,17,18,19]. Among e numerous integration systems, ΦC31 integrase has emerged as an outstanding one in many fields. us, we were inspired to develop an in vivo T7 integration system mediated by ΦC31 integrase. PhiC31 integrase, a site-specific large serine recombinase, is a useful tool for genome engineering in a variety of eukaryotic species and cell types. PhiC31 integrase performs efficient recombination between its attB site and ei er its own placed attP site or a partially mismatched genomic pseudo attP site. Home › About CIRM › Our Publications › Grantee publications › PhiC31 integrase for modification of stem cells PhiC31 integrase for modification of stem cells Journal.