Lois et al (2002) established a highly efficient method to generate transgenic animals with lentivirus. Lentivirus is injected to the space between a single cell embryio and its zona pellucida. In the host cell lentivirus genome will be reverese-transcribed and integrated into host genome randomly.
PiggyBac is a transposon isolated from cabbage looper moth Trichoplusia ni. PiggyBac transoson system works efficintly in mammlalian cells and has benn used in mouse genetics such as cancer genetics (Rad 2010) and random mutagenesis (Ding 2005, Sun 2008). PiggyBac has advantages over other transposon systems such as SleepingBeauty and Tol2 ;1) weak local hopping tendency, 2) large capacity, 3) clean "cut and paeste" hopping.
(A)Lentiviral probe. (B) PiggyBac probes.Our enhancer probe constructs use the tet-off system and incorporated a tet-responsive element driving the fluorescent reporter mCitrine, so that we could examine expression patterns in driver lines without crossing to separate reporter lines. tet-Cre probe expresses also Cre recombinase.
We established founder lines with single copy of PB insertion. Founders are crossed with trangenic animals with PBase. We used 2 PBase lines; Rosa-PBase (Rad 2010), which expressed mPBase nearly ubiquitously, and prm1-hyPBase in which protamine-1 promter drive hyper-active PBase (Yusa 2011) in spermatides.
- Dickson, J., Gowher, H., Strogantsev, R., Gaszner, M., Hair, A., Felsenfeld, G., & West, A. G. (2010). VEZF1 elements mediate protection from DNA methylation. PLoS Genetics, 6(1), e1000804. doi:10.1371/journal.pgen.1000804
- Ding, S., Wu, X., Li, G., Han, M., Zhuang, Y., & Xu, T. (2005). Efficient transposition of the piggyBac (PB) transposon in mammalian cells and mice. Cell, 122(3), 473–483. doi:10.1016/j.cell.2005.07.013
- Lois, C., Hong, E. J., Pease, S., Brown, E. J., & Baltimore, D. (2002). Germline Transmission and Tissue-Specific Expression of Transgenes Delivered by Lentiviral Vectors, 295(February), 868–872.
- Rad, R., Rad, L., Wang, W., Cadinanos, J., Vassiliou, G., Rice, S., et al. (2010). PiggyBac transposon mutagenesis: a tool for cancer gene discovery in mice. Science (New York, N.Y.), 330(6007), 1104–1107. doi:10.1126/science.1193004
- Sun, L. V., Jin, K., Liu, Y., Yang, W., Xie, X., Ye, L., et al. (2008). PBmice: an integrated database system of piggyBac (PB) insertional mutations and their characterizations in mice. Nucleic Acids Research, 36(Database issue), D729–34. http://doi.org/10.1093/nar/gkm790
- Yusa, K., Zhou, L., Li, M. A., Bradley, A., & Craig, N. L. (2011). A hyperactive piggyBac transposase for mammalian applications. Proceedings of the National Academy of Sciences, 108(4), 1531–1536. doi:10.1073/pnas.1008322108