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. Author manuscript; available in PMC: 2013 Oct 10.
Published in final edited form as: Virology. 2012 Jul 6;432(1):219–231. doi: 10.1016/j.virol.2012.06.007

Figure 4. Effects of CR8#13 on HIV-1 splicing.

Figure 4

A) Schematic of the pNLEnv plasmid. Envelope gene is expressed from an unspliced mRNA, whereas Nef is made from a spliced mRNA. B) 293T cells were transfected with pc-Tat plasmid (1 µg) and/or CR8#13 (10 nM). Western blot analysis using antibodies against Nef, or Env were performed after 3 days. Actin was used as control. To detect RNA levels of Nef and Env after drug treatment, 100 ng of total RNA was used for cDNA synthesis and PCR for Nef and Env regions. Total RNA samples were stained with ethidium bromide and run on a 2% denaturing gel. C) pNLEnv vector (20 µg) was transfected with pc-Tat (10 µg) into 293T cells and treated with CR8#13 (10 nM) for 3 days. Soluble nuclear proteins were obtained and 50 µg of nuclear extract was run on 4–20% SDS/PAGE and western blotted with α-Tat rabbit polyclonal antibody (5 µg; generous gift from Dr. Bryan Cullen; (Zhou et al., 2004).