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Please use this identifier to cite or link to this item: http://repository.li.mahidol.ac.th/dspace/handle/123456789/13690
Title: Germline transgenesis and insertional mutagenesis in Schistosoma mansoni mediated by murine leukemia virus
Authors: Gabriel Rinaldi
Sabine E. Eckert
Isheng J. Tsai
Sutas Suttiprapa
Kristine J. Kines
José F. Tort
Victoria H. Mann
Daniel J. Turner
Matthew Berriman
Paul J. Brindley
George Washington University
Universidad de la Republica Facultad de Medicina
Wellcome Trust Sanger Institute
Oxford Nanopore Technologies
Mahidol University
Tulane University School of Medicine
Keywords: Biochemistry, Genetics and Molecular Biology;Immunology and Microbiology
Issue Date: 1-Jul-2012
Citation: PLoS Pathogens. Vol.8, No.7 (2012), 16
Abstract: Functional studies will facilitate characterization of role and essentiality of newly available genome sequences of the human schistosomes, Schistosoma mansoni, S. japonicum and S. haematobium. To develop transgenesis as a functional approach for these pathogens, we previously demonstrated that pseudotyped murine leukemia virus (MLV) can transduce schistosomes leading to chromosomal integration of reporter transgenes and short hairpin RNA cassettes. Here we investigated vertical transmission of transgenes through the developmental cycle of S. mansoni after introducing transgenes into eggs. Although MLV infection of schistosome eggs from mouse livers was efficient in terms of snail infectivity, > 10-fold higher transgene copy numbers were detected in cercariae derived from in vitro laid eggs (IVLE). After infecting snails with miracidia from eggs transduced by MLV, sequencing of genomic DNA from cercariae released from the snails also revealed the presence of transgenes, demonstrating that transgenes had been transmitted through the asexual developmental cycle, and thereby confirming germline transgenesis. High-throughput sequencing of genomic DNA from schistosome populations exposed to MLV mapped widespread and random insertion of transgenes throughout the genome, along each of the autosomes and sex chromosomes, validating the utility of this approach for insertional mutagenesis. In addition, the germline-transmitted transgene encoding neomycin phosphotransferase rescued cultured schistosomules from toxicity of the antibiotic G418, and PCR analysis of eggs resulting from sexual reproduction of the transgenic worms in mice confirmed that retroviral transgenes were transmitted to the next (F1) generation. These findings provide the first description of wide-scale, random insertional mutagenesis of chromosomes and of germline transmission of a transgene in schistosomes. Transgenic lines of schistosomes expressing antibiotic resistance could advance functional genomics for these significant human pathogens. Database accession: Sequence data from this study have been submitted to the European Nucleotide Archive (http://www.ebi.ac.uk/embl) under accession number ERP000379. © 2012 Rinaldi et al.
URI: https://www.scopus.com/inward/record.uri?partnerID=HzOxMe3b&scp=84864591486&origin=inward
http://repository.li.mahidol.ac.th/dspace/handle/123456789/13690
ISSN: 15537374
15537366
Appears in Collections:Scopus 2011-2015

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