University Library
  • Login
A gateway to Melbourne's research publications
Minerva Access is the University's Institutional Repository. It aims to collect, preserve, and showcase the intellectual output of staff and students of the University of Melbourne for a global audience.
View Item 
  • Minerva Access
  • Medicine, Dentistry & Health Sciences
  • Melbourne Medical School
  • Medicine and Radiology
  • Medicine and Radiology - Research Publications
  • View Item
  • Minerva Access
  • Medicine, Dentistry & Health Sciences
  • Melbourne Medical School
  • Medicine and Radiology
  • Medicine and Radiology - Research Publications
  • View Item
JavaScript is disabled for your browser. Some features of this site may not work without it.

    Targeted insertion of an anti-CD2 monoclonal antibody transgene into the GGTA1 locus in pigs using FokI-dCas9

    Thumbnail
    Download
    Published version (1.406Mb)

    Citations
    Scopus
    Web of Science
    Altmetric
    13
    13
    Author
    Nottle, MB; Salvaris, EJ; Fisicaro, N; McIlfatrick, S; Vassiliev, I; Hawthorne, WJ; O'Connell, PJ; Brady, JL; Lew, AM; Cowan, PJ
    Date
    2017-08-16
    Source Title
    Scientific Reports
    Publisher
    NATURE PUBLISHING GROUP
    University of Melbourne Author/s
    Lew, Andrew; Salvaris, Evelyn; Cowan, Peter; Brady, Jamie
    Affiliation
    Medicine and Radiology
    Medical Biology (W.E.H.I.)
    Metadata
    Show full item record
    Document Type
    Journal Article
    Citations
    Nottle, M. B., Salvaris, E. J., Fisicaro, N., McIlfatrick, S., Vassiliev, I., Hawthorne, W. J., O'Connell, P. J., Brady, J. L., Lew, A. M. & Cowan, P. J. (2017). Targeted insertion of an anti-CD2 monoclonal antibody transgene into the GGTA1 locus in pigs using FokI-dCas9. SCIENTIFIC REPORTS, 7 (1), https://doi.org/10.1038/s41598-017-09030-6.
    Access Status
    Open Access
    URI
    http://hdl.handle.net/11343/254930
    DOI
    10.1038/s41598-017-09030-6
    Abstract
    Xenotransplantation from pigs has been advocated as a solution to the perennial shortage of donated human organs and tissues. CRISPR/Cas9 has facilitated the silencing of genes in donor pigs that contribute to xenograft rejection. However, the generation of modified pigs using second-generation nucleases with much lower off-target mutation rates than Cas9, such as FokI-dCas9, has not been reported. Furthermore, there have been no reports on the use of CRISPR to knock protective transgenes into detrimental porcine genes. In this study, we used FokI-dCas9 with two guide RNAs to integrate a 7.1 kilobase pair transgene into exon 9 of the GGTA1 gene in porcine fetal fibroblasts. The modified cells lacked expression of the αGal xenoantigen, and secreted an anti-CD2 monoclonal antibody encoded by the transgene. PCR and sequencing revealed precise integration of the transgene into one allele of GGTA1, and a small deletion in the second allele. The cells were used for somatic cell nuclear transfer to generate healthy male knock-in piglets, which did not express αGal and which contained anti-CD2 in their serum. We have therefore developed a versatile high-fidelity system for knocking transgenes into the pig genome for xenotransplantation purposes.

    Export Reference in RIS Format     

    Endnote

    • Click on "Export Reference in RIS Format" and choose "open with... Endnote".

    Refworks

    • Click on "Export Reference in RIS Format". Login to Refworks, go to References => Import References


    Collections
    • Minerva Elements Records [45770]
    • Medical Biology - Research Publications [865]
    • Medicine and Radiology - Research Publications [2347]
    Minerva AccessDepositing Your Work (for University of Melbourne Staff and Students)NewsFAQs

    BrowseCommunities & CollectionsBy Issue DateAuthorsTitlesSubjectsThis CollectionBy Issue DateAuthorsTitlesSubjects
    My AccountLoginRegister
    StatisticsMost Popular ItemsStatistics by CountryMost Popular Authors