Fig. 3

The R2Ol system generates transgenic zebrafish. a (1)–(3) Schematic diagram of R2Ol with transgene cassettes used in the retrotransposition assay. EGFP cassette includes EF1, Xenopus EF1-alpha promoter; EGFP, enhanced green fluorescent protein (EGFP) ORF forward or reverse orientations; PA, SV40 poly(A) signal; GAL4 cassette includes heat shock promoter (hsp), and GAL4 transcription factor. All constructs contain a 105-bp 28S rDNA sequence upstream of the 5′ UTR to increase full-length insertion and a 10-bp 28S rDNA sequence downstream of the 3′ UTR. (4) Two constructs for the trans-complementation assay. F0 retrotransposition efficiency and founder efficiency for each construct are indicated on the right (see also Table 1). b F0 founder screening and off target detection. Top, PCR founder screening of the 3′ junction region using pooled F1 genomic DNA. Primers are indicated by arrows. Bottom, internal PCR to detect external 28S rDNA insertions. Red asterisks indicate the obtained founders used in further analyses. Blue asterisks indicate internal PCR results