ArticleNature protocols2014
Germline transgenesis in pigs by cytoplasmic microinjection of Sleeping Beauty transposons.
Article in Nature protocols, 2014. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 37 papers.
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Who cites it
37 citing papers in PubMed, 71 citations in OpenAlex.
- Induced Pluripotent Stem Cells in Birds: Opportunities and Challenges for Science and Agriculture.Veterinary sciences · 2024Review
- A versatile laser-induced porcine model of outer retinal and choroidal degeneration for preclinical testing.JCI insight · 2023Article
- Multiple gene editing in porcine embryos using a combination of microinjection, electroporation, and transfection methods.Veterinary world · 2022Article
- Contemporary Transposon Tools: A Review and Guide through Mechanisms and Applications ofInternational journal of molecular sciences · 2021Review
- One-step genome editing of porcine zygotes through the electroporation of a CRISPR/Cas9 system with two guide RNAs.In vitro cellular & developmental biology. Animal · 2020Article
- A single amino acid switch converts the Sleeping Beauty transposase into an efficient unidirectional excisionase with utility in stem cell reprogramming.Nucleic acids research · 2020Article
- Optimization Strategy for Generating Gene-edited Tibet Minipigs by Synchronized Oestrus and Cytoplasmic Microinjection.International journal of biological sciences · 2019Article
- Enhancer Trapping and Annotation in Zebrafish Mediated with Sleeping Beauty, piggyBac and Tol2 Transposons.Genes · 2018Article
- Changes in Skeletal Muscle and Body Weight on Sleeping Beauty Transposon-Mediated Transgenic Mice Overexpressing Pig mIGF-1.Biochemical genetics · 2018Article
- Long-term health and germline transmission in transgenic cattle following transposon-mediated gene transfer.BMC genomics · 2018Article
- Efficient Non-viral Gene Delivery into Human Hematopoietic Stem Cells by Minicircle Sleeping Beauty Transposon Vectors.Molecular therapy : the journal of the American Society of Gene Therapy · 2018Article
- Genetically engineered pigs as models for human disease.Disease models & mechanisms · 2018Review
- Cell-penetrating peptide-driven Cre recombination in porcine primary cells and generation of marker-free pigs.PloS one · 2018Article
- Tracking mesenchymal stem cell contributions to regeneration in an immunocompetent cartilage regeneration model.JCI insight · 2017Article
- RNAi combining Sleeping Beauty transposon system inhibits ex vivo expression of foot-and-mouth disease virus VP1 in transgenic sheep cells.Scientific reports · 2017Article
- A vector platform for the rapid and efficient engineering of stable complex transgenes.Scientific reports · 2016Article
- Characterization of Growth and Reproduction Performance, Transgene Integration, Expression, and Transmission Patterns in Transgenic Pigs Produced by piggyBac Transposition-Mediated Gene Transfer.Animal biotechnology · 2016Article
- Factors affecting efficiency of introducing foreign DNA and RNA into parthenogenetic or in vitro-fertilized porcine eggs by cytoplasmic microinjection.In vitro cellular & developmental biology. Animal · 2016Article
- Engineering Large Animal Species to Model Human Diseases.Current protocols in human genetics · 2016Article
- Gene targeting, genome editing: from Dolly to editors.Transgenic research · 2016Review
Corrections and comments
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Authors and funding
12 authors at 7 institutions in 5 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
The pig has emerged as an important large animal model in biomedical and pharmaceutical research. We describe a protocol for high-efficiency germline transgenesis and sustained transgene expression in pigs by using the Sleeping Beauty (SB) transposon system. The protocol is based on co-injection of a plasmid encoding the SB100X hyperactive transposase, together with a second plasmid carrying a transgene flanked by binding sites for the transposase, into the cytoplasm of porcine zygotes. The transposase mediates excision of the transgene cassette from the plasmid vector and its permanent insertion into the genome to produce stable transgenic animals. This method compares favorably in terms of both efficiency and reliable transgene expression to classic pronuclear microinjection or somatic cell nuclear transfer (SCNT), and it offers comparable efficacies to lentiviral approaches, without limitations on vector design, issues of transgene silencing and the toxicity and biosafety concerns of working with viral vectors. Microinjection of the vectors into zygotes and transfer of the embryos to recipient animals can be performed in 1 d; generation of germline-transgenic lines by using this protocol takes ∼1 year.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.