Lentiviral vectors: are they the future of animal transgenesis?

Park, Frank

doi:10.1152/physiolgenomics.00069.2007

Cited by 147 publications

(118 citation statements)

References 142 publications

(244 reference statements)

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“…Furthermore, unlike retroviral vectors [43][44][45][46] , SB100X transposase-catalyzed transgene integration does not seem to trigger transcriptional silencing 34,39 . Importantly, SB transposon-tagged genomic loci are suitable for recombinase-mediated cassette exchange (RMCE) 47 , allowing targeted genome engineering 5,42 .…”

Section: Transgenesis Using the Sleeping Beauty Transposonmentioning

confidence: 98%

Germline transgenesis in pigs by cytoplasmic microinjection of Sleeping Beauty transposons

et al. 2014

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2The 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 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, and offers comparable efficacies to lentiviral approaches, without limitations on vector design, issues of transgene silencing as well as 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 one day; generation of germline-transgenic lines by using this protocol takes approximately one year.

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Section: Transgenesis Using the Sleeping Beauty Transposonmentioning

confidence: 98%

Germline transgenesis in pigs by cytoplasmic microinjection of Sleeping Beauty transposons

et al. 2014

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“…Furthermore, unlike retroviral vectors [34][35][36][37] , SB100X transposase-catalyzed transgene integration does not seem to trigger transcriptional silencing 24,30 . Therefore, the application of the Sleeping Beauty transposon system described here can significantly enhance the rabbit genomic toolbox.…”

Section: Transgenesis With the Sleeping Beauty Transposonmentioning

confidence: 98%

Germline transgenesis in rabbits by pronuclear microinjection of Sleeping Beauty transposons

et al. 2014

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2The laboratory rabbit (Oryctolagus cuniculus) is widely used as a model for a variety of inherited and acquired human diseases. In addition, the rabbit is the smallest livestock animal that is used to transgenically produce pharmaceutical proteins in its milk. Here we describe a protocol for highefficiency germline transgenesis and sustained transgene expression in rabbits by using the Sleeping Beauty transposon system. The protocol is based on co-injection into the pronuclei of fertilized oocytes of synthetic mRNA encoding the SB100X hyperactive transposase, together with plasmid DNA carrying a transgene construct flanked by binding sites for the transposase. The translation of the transposase mRNA is followed by enzyme-mediated excision of the transgene cassette from the plasmids and its permanent genomic insertion to produce stable transgenic animals. Generation of a germline-transgenic founder animal by using this protocol takes approximately two months.Transposon-mediated transgenesis compares favorably in terms of both efficiency and reliable transgene expression to classic pronuclear microinjection, and offers comparable efficacies (numbers of transgenic founders obtained per injected embryo) to lentiviral approaches, without limitations on vector design, issues of transgene silencing as well as the toxicity and biosafety concerns of working with viral vectors.

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“…This high expression occurs because the integration of the lentivirus occurs preferentially in active transcriptional units in the host genome. 20 The use of plasmid vectors based on transposons system has emerged as an attractive alternative for generating transgenic animals. Sleeping Beauty, piggyBac and Tol2 transposons have been developed as gene transfer methods for vertebrate genetics, and commercial vectors are available (Addgene).…”

Section: Vectors Used In the Generation Of Transgenic Animalsmentioning

confidence: 99%

Transgenic bovine as bioreactors: Challenges and perspectives

et al. 2016

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The use of recombinant proteins has increased in diverse commercial sectors. Various systems for protein production have been used for the optimization of production and functional protein expression. The mammary gland is considered to be a very interesting system for the production of recombinant proteins due to its high level of expression and its ability to perform post-translational modifications. Cows produce large quantities of milk over a long period of lactation, and therefore this species is an important candidate for recombinant protein expression in milk. However, transgenic cows are more difficult to generate due to the inefficiency of transgenic methodologies, the long periods for transgene detection, recombinant protein expression and the fact that only a single calf is obtained at the end of each pregnancy. An increase in efficiency for transgenic methodologies for cattle is a big challenge to overcome. Promising methodologies have been proposed that can help to overcome this obstacle, enabling the use of transgenic cattle as bioreactors for protein production in milk for industry.

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Lentiviral vectors: are they the future of animal transgenesis?

Cited by 147 publications

References 142 publications

Germline transgenesis in pigs by cytoplasmic microinjection of Sleeping Beauty transposons

Germline transgenesis in pigs by cytoplasmic microinjection of Sleeping Beauty transposons

Germline transgenesis in rabbits by pronuclear microinjection of Sleeping Beauty transposons

Transgenic bovine as bioreactors: Challenges and perspectives

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