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. Author manuscript; available in PMC: 2019 Jul 1.
Published in final edited form as: Virology. 2018 May 29;520:116–126. doi: 10.1016/j.virol.2018.05.016

Fig. 5: Efficient introduction of indels into unintegrated HIV-1 proviruses after Cas9 cleavage.

Fig. 5:

(A) The level of indel introduction into TAR was assayed as described in Fig. 2 by Sacl cleavage of PCR-amplified TAR sequences recovered from 293T cells expressing Sp Cas9 and the TAR1 and TAR2 sgRNAs in the presence or absence of Raltegravir. (B) Production of NLuc by the infecting NL-NLuc-HXB indicator provirus at 72 h post-infection in cells expressing Sp Cas9 and either a control sgRNA or both the TAR1 and TAR2 sgRNAs in the presence or absence of Raltegravir. Average of three independent experiments with SD indicated. (C) HIV-1 LTRs bearing indels in TAR are largely transcriptionally inactive. RT-PCR was used to amplify DNA encoding TAR from viral mRNAs expressed in HIV-1 infected cells expressing Sp Cas9 and the TAR1 and TAR2 sgRNAs, as shown in panel A. This cDNA was then analyzed for the presence of TAR indels by SacI cleavage, as described in Fig. 2.