Efficient generation of transgenic mice by lentivirus‐mediated modification of spermatozoa

The FASEB JournalVolume 28, Issue 2 p. 569-576 HypothesesFree to Read Efficient generation of transgenic mice by lentivirus-mediated modification of spermatozoa Anil Chandrashekran, Corresponding Author Anil Chandrashekran achandrashekran@rvc.ac.uk Department of Surgery and Cancer, Division of Cancer, Institute of Reproductive and Developmental Biology (IRDB), Imperial College London, London, UKCorrespondence and current address: Department of Veterinary Clinical Sciences, The Royal Veterinary College, Hawkshead Ln., North Mimms, AL9 7TA, UK. E-mail: achandrashekran@rvc.ac.ukSearch for more papers by this authorRupa Sarkar, Rupa Sarkar Department of Surgery and Cancer, Division of Cancer, Institute of Reproductive and Developmental Biology (IRDB), Imperial College London, London, UKSearch for more papers by this authorAdrian Thrasher, Adrian Thrasher Molecular Immunology Unit, University College London Institute of Child Health, London, UKSearch for more papers by this authorScott E. Fraser, Scott E. Fraser Biological Imaging Center, Beckman Institute, California Institute of Technology, Pasadena, California, USASearch for more papers by this authorNicholas Dibb, Nicholas Dibb Department of Surgery and Cancer, Division of Cancer, Institute of Reproductive and Developmental Biology (IRDB), Imperial College London, London, UKSearch for more papers by this authorColin Casimir, Colin Casimir Department of Natural Sciences, School of Science and Technology, Middlesex University, London, UKSearch for more papers by this authorRobert Winston, Robert Winston Department of Surgery and Cancer, Division of Cancer, Institute of Reproductive and Developmental Biology (IRDB), Imperial College London, London, UKSearch for more papers by this authorCarol Readhead, Carol Readhead Biological Imaging Center, Beckman Institute, California Institute of Technology, Pasadena, California, USASearch for more papers by this author Anil Chandrashekran, Corresponding Author Anil Chandrashekran achandrashekran@rvc.ac.uk Department of Surgery and Cancer, Division of Cancer, Institute of Reproductive and Developmental Biology (IRDB), Imperial College London, London, UKCorrespondence and current address: Department of Veterinary Clinical Sciences, The Royal Veterinary College, Hawkshead Ln., North Mimms, AL9 7TA, UK. E-mail: achandrashekran@rvc.ac.ukSearch for more papers by this authorRupa Sarkar, Rupa Sarkar Department of Surgery and Cancer, Division of Cancer, Institute of Reproductive and Developmental Biology (IRDB), Imperial College London, London, UKSearch for more papers by this authorAdrian Thrasher, Adrian Thrasher Molecular Immunology Unit, University College London Institute of Child Health, London, UKSearch for more papers by this authorScott E. Fraser, Scott E. Fraser Biological Imaging Center, Beckman Institute, California Institute of Technology, Pasadena, California, USASearch for more papers by this authorNicholas Dibb, Nicholas Dibb Department of Surgery and Cancer, Division of Cancer, Institute of Reproductive and Developmental Biology (IRDB), Imperial College London, London, UKSearch for more papers by this authorColin Casimir, Colin Casimir Department of Natural Sciences, School of Science and Technology, Middlesex University, London, UKSearch for more papers by this authorRobert Winston, Robert Winston Department of Surgery and Cancer, Division of Cancer, Institute of Reproductive and Developmental Biology (IRDB), Imperial College London, London, UKSearch for more papers by this authorCarol Readhead, Carol Readhead Biological Imaging Center, Beckman Institute, California Institute of Technology, Pasadena, California, USASearch for more papers by this author First published: 02 December 2013 https://doi.org/10.1096/fj.13-233999Citations: 12 This article includes supplemental data. Please visit http://www.fasebj.org to obtain this information. Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat Abstract Transgenic technologies conventionally rely on the oocyte as a substrate for genetic modification. Owing to their accessibility, however, male germ cells, including mature sperm, have material advantages for use in transgenesis. Here we have exploited lentiviruses to generate transgenic animals via the male germline. When pseudotyped lentiviral vectors encoding green fluorescent protein (GFP) were incubated with mouse spermatozoa, these sperm were highly successful in producing transgenics. Lentivirally transduced mouse spermatozoa were used in in vitro fertilization (IVF) studies, and when followed by embryo transfer, ≥42% of founders were found to be transgenic for GFP. Inverse PCR strategy for integration site analysis demonstrated integration of at least 1 or 2 copies of GFP in the transgenics, mapping to different chromosomes. GFP expression was detected in a wide range of murine tissues, including testis and the transgene was stably transmitted to a third generation of transgenic animals. This relatively simple, yet highly efficient, technique for generating transgenic animals by transducing spermatozoa with lentiviral vectors in vitro is a powerful tool for the study of fertilization/preimplantation development, vertical viral gene transmission, gene function and regulation, and epigenetic inheritance.—Chandrashekran, A., Sarkar, R., Thrasher, A., Fraser, S. E., Dibb, N., Casimir, C., Readhead, C., Winston, R. Efficient generation of transgenic mice by lentivirus-mediated modification of spermatozoa. FASEB J. 28, 569–576 (2014). www.fasebj.org Citing Literature Supporting Information Filename Description fsb2028002005-sup-0001.zipZip archive, 4.5 MB Supplementary material Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article. Volume28, Issue2February 2014Pages 569-576 RelatedInformation