Mutational analysis of IΔ1 type IAP element. (A) Western blot analysis of the HA-tagged IΔ1 type IAP protein. pDE1ex-HA contains HA tag at the C terminus of gag-pol open reading frame. Expected size (∼125 kDa) was observed (indicated by the arrowhead). Mock, pBluescriptII; Actb, beta-actin. (B) Effect of mutation in Gag region on trans-complementation. pFL, pDE1, and pDE1-M3 were transfected into HeLa cells with (+) or without (−) pFLex, which expresses autonomous type IAP protein, and transposition efficiency was examined by GFP signal. Data are mean ± SEM of three independent experiments. ††P < 0.01 (t-test). (C) Real-time RT-PCR analysis of the expression levels of IAP transcripts from wild-type and mutated vectors. Data are mean ± SEM of three independent experiments. (D) Northern blot analysis of the expression levels of IAP transcripts. Arrowhead indicates bands with predicted size; asterisk denotes unpredicted bands.
