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. 1999 Nov 9;96(23):13118–13123. doi: 10.1073/pnas.96.23.13118

Figure 6.

Figure 6

Test for IRES activity in the 5′ NCRs of Pim-1 and Cyr61 mRNAs. (A) Expression of dicistronic mRNAs containing ΔEMCV sequences or the 5′NCRs of Cyr61 or Pim-1 in the intercistronic spacer regions. Plasmid DNA (5 μg) and 25 μg of lipofectin reagent (GIBCO/BRL) were mixed and transfected into HeLa cells. The results are displayed as the ratio of second cistron fLuc to first cistron rLuc activity. The ratios of luciferase activities obtained from transfection experiments with the control plasmid pRΔDEF was set to 1. Results from three independent transfections were averaged and are shown. (B) Integrity of dicistronic mRNAs in transfected cells. Poly(A+) mRNA was isolated from cells transfected with dicistronic plasmids and was analyzed by Northern blot hybridization using radiolabeled DNA probes complementary to firefly luciferase sequences. The migrations of 28S and 18S rRNA are indicated.