Figure 4—figure supplement 2. | RNA-directed remodeling of the HIV-1 protein Rev orchestrates assembly of the Rev–Rev response element complex

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RNA-directed remodeling of the HIV-1 protein Rev orchestrates assembly of the Rev–Rev response element complex

Figure 4—figure supplement 2.

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University of California, San Francisco, United States
Figure 4—figure supplement 2.
Download figureOpen in new tabFigure 4—figure supplement 2. Design of virus constructs.

The endogenous rev locus was deleted by mutating the start codon (ATG) to ACG and introducing a termination codon at residue 23. Both mutations are silent in the overlapping tat open reading frame. The nef locus, dispensable for virus replication ex vivo, was disrupted by introducing SacII and Xba1 restriction endonuclease sites in place of the start ATG codon. Codon-optimized Rev DNA was synthesized (to avoid the introduction of direct nucleotide repeats within the HIV genome that typically results in recombination) with the endogenous Kozak sequence and flanking SacII and XbaI restriction sites to facilitate cloning within the repurposed nef locus.

DOI: http://dx.doi.org/10.7554/eLife.04120.012