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. 2004 Sep;24(18):7902–7913. doi: 10.1128/MCB.24.18.7902-7913.2004

FIG. 6.

FIG. 6.

The EGFP gene connected with the SART1 3′ UTR is retrotransposed into the telomeric repeats. (A) Schematic of trans complementation between EGFP-S1/3′UTR (a) and Δ3′UTR (b). EGFP1/S1-3′UTR is an AcNPV-expressed clone that includes the EGFP gene directly followed by the complete SART1 3′ UTR. Δ3′UTR does not have retrotransposition activity because it lacks the 3′ UTR but can provide the ORF1-ORF2 protein complex. 2D699V is a mutant that has lost its RT activity (24). The 3′ junction of in vivo retrotransposed copies was detected by PCR with primers EGFP1 S688 and (CCTAA)6 (shown by white arrows), and the 5′ junction was detected by PCR with primers EGFP1 A576 and (TTAGG)6 (shown by black arrows). APE, apurinic/apyrimidinic EN. (B) Ethidium bromide-stained gels of the junction PCR assay for in vivo retrotransposition assay. Left, 3′ junction PCR; right, 5′ junction PCR; a, EGFP1/S1-3′UTR infection of Sf9 cells; a+b, coinfection with EGFP1/S1-3′UTR and Δ3′UTR; a+c, coinfection with EGFP1/S1-3′UTR and 2D699V; Sf9, cells without AcNPV infection. The black arrowhead indicates the successful in vivo retrotransposition band with the expected size. Molecular sizes are shown on the left.