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Transgenic rabbit production with simian immunodeficiency virus-derived lentiviral vector

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Abstract

Transgenic rabbit is the preferred disease model of atherosclerosis, lipoprotein metabolism and cardiovascular diseases since upon introducing genetic mutations of human genes, rabbit models reflect human physiological and pathological states more accurately than mouse models. Beyond that, transgenic rabbits are also used as bioreactors to produce pharmaceutical proteins in their milk. Since in the laboratory rabbit the conventional transgenesis has worked with the same low efficiency in the last twenty five years and truly pluripotent embryonic stem cells are not available to perform targeted mutagenesis, our aim was to adapt lentiviral transgenesis to this species. A simian immunodeficiency virus based replication defective lentiviral vector was used to create transgenic rabbit through perivitelline space injection of fertilized oocytes. The enhanced green fluorescent protein (GFP) gene was placed under the ubiquitous CAG promoter. Transgenic founder rabbits showed mosaic pattern of GFP expression. Transgene integration and expression was revealed in tissues derived from all three primary germ layers. Transgene expression was detected in the developing sperm cells and could get through the germ line without epigenetic silencing, albeit with very low frequency. Our data show for the first time, that lentiviral transgenesis could be a feasible and viable alternative method to create genetically modified laboratory rabbit.

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References

  • Besenfelder U, Aigner B, Müller M, Brem G (1998) Generation and application of transgenic rabbits. In: Cid-Arregui A, Garcia-Carrancá A (eds) Microinjection and transgenesis: strategies and protocol. Springer-Verlag, Berlin, pp 561–586

  • Bodrogi L, Brands R, Raaben W, Seinen W, Baranyi M, Fiechter D, Bosze Z (2006) High level expression of tissue-nonspecific alkaline phosphatase in the milk of transgenic rabbits. Transgenic Res 15(5):627–636

    Article  CAS  PubMed  Google Scholar 

  • Bosze Z, Houdebine L-M (2006) Application of rabbits in biomedical research: a review. World Rabbit Sci 14:1–14

    Google Scholar 

  • Cutino-Moguel T, Fassati A (2006) A phenotypic recessive, post-entry block in rabbit cells that results in aberrant trafficking of HIV-1. Traffic 7(8):978–992

    Article  CAS  PubMed  Google Scholar 

  • Derdouch S, Gay W, Negre D, Prost S, Le Dantec M, Delache B, Auregan G, Andrieu T, Leplat JJ, Cosset FL, Le Grand R (2008) Reconstitution of the myeloid and lymphoid compartments after the transplantation of autologous and genetically modified CD34+ bone marrow cells, following gamma irradiation in cynomolgus macaques. Retrovirology 5:50

    Article  PubMed  Google Scholar 

  • Graur D, Duret L, Gouy M (1996) Phylogenetic position of the order Lagomorpha (rabbits, hares and allies). Nature 379(6563):333–335

    Article  CAS  PubMed  Google Scholar 

  • Hammer RE, Pursel VG, Rexroad CE Jr, Wall RJ, Bolt DJ, Ebert KM, Palmiter RD, Brinster RL (1985) Production of transgenic rabbits, sheep and pigs by microinjection. Nature 315(6021):680–683

    Article  CAS  PubMed  Google Scholar 

  • Hiripi L, Makovics F, Halter R, Baranyi M, Paul D, Carnwath JW, Bosze Z, Niemann H (2003) Expression of active human blood clotting factor VIII in mammary gland of transgenic rabbits. DNA Cell Biol 22(1):41–45

    Article  CAS  PubMed  Google Scholar 

  • Hofmann W, Schubert D, LaBonte J, Munson L, Gibson S, Scammell J, Ferrigno P, Sodroski J (1999) Species-specific, postentry barriers to primate immunodeficiency virus infection. J Virol 73(12):10020–10028

    CAS  PubMed  Google Scholar 

  • Hofmann A, Kessler B, Ewerling S, Weppert M, Vogg B, Ludwig H, Stojkovic M, Boelhauve M, Brem G, Wolf E, Pfeifer A (2003) Efficient transgenesis in farm animals by lentiviral vectors. EMBO Rep 4(11):1054–1060

    Article  CAS  PubMed  Google Scholar 

  • Hofmann A, Zakhartchenko V, Weppert M, Sebald H, Wenigerkind H, Brem G, Wolf E, Pfeifer A (2004) Generation of transgenic cattle by lentiviral gene transfer into oocytes. Biol Reprod 71(2):405–409

    Article  CAS  PubMed  Google Scholar 

  • Hofmann A, Kessler B, Ewerling S, Kabermann A, Brem G, Wolf E, Pfeifer A (2006) Epigenetic regulation of lentiviral transgene vectors in a large animal model. Mol Ther 13(1):59–66

    Article  CAS  PubMed  Google Scholar 

  • Kvell K, Nguyen TH, Salmon P, Glauser F, Werner-Favre C, Barnet M, Schneider P, Trono D, Zubler RH (2005) Transduction of CpG DNA-stimulated primary human B cells with bicistronic lentivectors. Mol Ther 12(5):892–899

    Article  CAS  PubMed  Google Scholar 

  • Kvell K, Czompoly T, Hiripi L, Balogh P, Kobor J, Bodrogi L, Pongracz JE, Ritchie WA, Bosze Z (2010) Characterisation of eGFP-transgenic BALB/c mouse strain established by lentiviral transgenesis. Transgenic Res 19(1):105–112

    Google Scholar 

  • Lois C, Hong EJ, Pease S, Brown EJ, Baltimore D (2002) Germline transmission and tissue-specific expression of transgenes delivered by lentiviral vectors. Science 295(5556):868–872

    Article  CAS  PubMed  Google Scholar 

  • McGrew MJ, Sherman A, Ellard FM, Lillico SG, Gilhooley HJ, Kingsman AJ, Mitrophanous KA, Sang H (2004) Efficient production of germline transgenic chickens using lentiviral vectors. EMBO Rep 5(7):728–733

    Article  CAS  PubMed  Google Scholar 

  • Naldini L, Blomer U, Gallay P, Ory D, Mulligan R, Gage FH, Verma IM, Trono D (1996) In vivo gene delivery and stable transduction of nondividing cells by a lentiviral vector. Science 272(5259):263–267

    Article  CAS  PubMed  Google Scholar 

  • Negre D, Cosset FL (2002) Vectors derived from simian immunodeficiency virus (SIV). Biochimie 84(11):1161–1171

    Article  CAS  PubMed  Google Scholar 

  • Negre D, Mangeot PE, Duisit G, Blanchard S, Vidalain PO, Leissner P, Winter AJ, Rabourdin-Combe C, Mehtali M, Moullier P, Darlix JL, Cosset FL (2000) Characterization of novel safe lentiviral vectors derived from simian immunodeficiency virus (SIVmac251) that efficiently transduce mature human dendritic cells. Gene Ther 7(19):1613–1623

    Article  CAS  PubMed  Google Scholar 

  • Park F (2007) Lentiviral vectors: are they the future of animal transgenesis? Physiol Genomics 31(2):159–173

    Article  CAS  PubMed  Google Scholar 

  • Pfeifer A, Ikawa M, Dayn Y, Verma IM (2002) Transgenesis by lentiviral vectors: lack of gene silencing in mammalian embryonic stem cells and preimplantation embryos. Proc Natl Acad Sci USA 99(4):2140–2145

    Article  CAS  PubMed  Google Scholar 

  • Ritchie WA, Neil C, King T, Whitelaw CB (2007) Transgenic embryos and mice produced from low titre lentiviral vectors. Transgenic Res 16(5):661–664

    Article  CAS  PubMed  Google Scholar 

  • Sasaki E, Suemizu H, Shimada A, Hanazawa K, Oiwa R, Kamioka M, Tomioka I, Sotomaru Y, Hirakawa R, Eto T, Shiozawa S, Maeda T, Ito M, Ito R, Kito C, Yagihashi C, Kawai K, Miyoshi H, Tanioka Y, Tamaoki N, Habu S, Okano H, Nomura T (2009) Generation of transgenic non-human primates with germline transmission. Nature 459(7246):523–U550

    Article  CAS  PubMed  Google Scholar 

  • Shen W, Li L, Pan Q, Min L, Dong H, Deng J (2006) Efficient and simple production of transgenic mice and rabbits using the new DMSO-sperm mediated exogenous DNA transfer method. Mol Reprod Dev 73(5):589–594

    Article  CAS  PubMed  Google Scholar 

  • Sultana F, Hatori M, Shimozawa N, Ebisawa T, Sankai T (2009) Continuous observation of rabbit preimplantation embryos in vitro by using a culture device connected to a microscope. J Am Assoc Lab Anim Sci 48(1):52–56

    CAS  PubMed  Google Scholar 

  • Takahashi R, Kuramochi T, Aoyagi K, Hashimoto S, Miyoshi I, Kasai N, Hakamata Y, Kobayashi E, Ueda M (2007) Establishment and characterization of CAG/EGFP transgenic rabbit line. Transgenic Res 16(1):115–120

    Article  CAS  PubMed  Google Scholar 

  • Whitelaw CB, Radcliffe PA, Ritchie WA, Carlisle A, Ellard FM, Pena RN, Rowe J, Clark AJ, King TJ, Mitrophanous KA (2004) Efficient generation of transgenic pigs using equine infectious anaemia virus (EIAV) derived vector. FEBS Lett 571(1–3):233–236

    Article  CAS  PubMed  Google Scholar 

  • Witschi E (1956) Development of vertebrates. Saunders W.B, Philadelphia

    Google Scholar 

  • Wolfgang MJ, Eisele SG, Browne MA, Schotzko ML, Garthwaite MA, Durning M, Ramezani A, Hawley RG, Thomson JA, Golos TG (2001) Rhesus monkey placental transgene expression after lentiviral gene transfer into preimplantation embryos. Proc Natl Acad Sci USA 98(19):10728–10732

    Article  CAS  PubMed  Google Scholar 

  • Zaitseva L, Cherepanov P, Leyens L, Wilson SJ, Rasaiyaah J, Fassati A (2009) HIV-1 exploits importin 7 to maximize nuclear import of its DNA genome. Retrovirology 6:11

    Article  PubMed  Google Scholar 

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Acknowledgments

We thank Ariberto Fassati (University College, London, UK), Bruce Whitelaw and Bill Ritchie (Roslin, UK) and Marielle Afanassieff (INRA, Lyon, France) for the advices and helpful discussions. The authors thank G. Takács for the artwork. Supporting grants: OTKA T049034, GVOP-3.1.1.-2004-05-0071/3.0 and OM-00118/2008, OTKA PD78310.

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Correspondence to Zs. Bősze.

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11248_2009_9356_MOESM1_ESM.tif

Supplementary Fig. 1. Integrated transgene copy number determination with Southern analysis. (A) Southern blot of BamHI-digested genomic DNA isolated from SIV-CAG-eGFP transgenic founder rabbit ear samples. Lines: 1–2: founder rabbit; line 3: non-transgenic rabbit, negative control; line 4: GFP-transgenic mouse (Kvell et al. 2009), positive control. (B) The lentiviral vector carrying the CAG-GFP transgene, LTR, long terminal repeat; CPPT, polypurine tract; WPRE, woodchuck hepatitis responsive element; dotted lines, rabbit genome. (TIFF 1652 kb)

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Hiripi, L., Negre, D., Cosset, FL. et al. Transgenic rabbit production with simian immunodeficiency virus-derived lentiviral vector. Transgenic Res 19, 799–808 (2010). https://doi.org/10.1007/s11248-009-9356-y

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  • DOI: https://doi.org/10.1007/s11248-009-9356-y

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