Transgene expression in zebrafish: A comparison of retroviral-vector and DNA-injection approaches

To assess alternative methods for introducing expressing transgenes into th e germ line of zebrafish, transgenic fish that express a nuclear-targeted, enhanced, green fluorescent protein (eGFP) gene were produced using both ps eudotyped retroviral vector infection and DNA microinjection of embryos. Ge rm-line transgenic founders were identified and the embryonic progeny of th ese founders were evaluated for the extent and pattern of eGFP expression. To compare the two modes of transgenesis, both vectors used the Xenopus tra nslational elongation factor 1-alpha enhancer/promoter regulatory cassette. Several transgenic founder fish which transferred eGFP expression to their progeny were identified. The gene expression patterns are described and co mpared for the two modes of gene transfer. Transient expression of eGFP was detected 1 day after introducing the transgenes via either DNA microinject ion or retroviral vector infection. In both cases of gene transfer, transge nic females produced eGFP-positive progeny even before the zygotic genome w as turned on. Therefore, GFP was being provided by the oocyte before fertil ization. A transgenic female revealed eGFP expression in her ovarian follic les. The qualitative patterns of gene expression in the transgenic progeny embryos after zygotic induction of gene expression were similar and indepen dent of the mode of transgenesis. The appearance of newly synthesized GEP i s detectable within 5-7 h after fertilization. The variability of the exten t of eGFP expression from transgenic founder to transgenic founder was wide r for the DNA-injection transgenics than for the retroviral vector-produced transgenics. The ability to provide expressing germ-line transgenic progen y via retroviral vector infection provides both an alternative mode of tran sgenesis for zebrafish work and a possible means of easily assessing the in sertional mutagenesis frequency of retroviral vector infection of zebrafish embryos. However, because of the transfer of GFP from oocyte to embryo, th e stability of GFP may create problems of analysis in embryos which develop as quickly as those of zebrafish. (C) 1999 Academic Press.