CRISPR/Cas9 and HIV
CRISPRs (clustered regularly interspaced short palindromic repeats) are regions where Bacteria and Archea incorporate foreign (plasmids, bacteriophage) genetic material into their own genome. If the bacteria encounters a similar foreign piece of DNA in the future, it will destroy it with enzymes. CRISPR RNA can be guided to particular gene sequences recognized by guide RNA. If the guide RNA complements the DNA, it starts cutting with Cas9 endonuclease. Recently, this technology has been used in several other models, including plants, mice, and even human embryos. The possibilities are amazing, as Wang et al. demonstrate in using a CRISPR/Cas9 system in their study of HIV resistance.
Adapted from: Wang et al. 2014. PLOS ONE. 9:e115987. Pubmed
Because R5-tropic HIV-1 is typically associated with sexual transmission, maternal-infant exposure, and percutaneous inoculation, CCR5 has been a focal point for disruption. Wang et al. utilized a single guided RNA (sgRNA) to regions of CCR5 (CR2) and transducted a T cell line with Cas9 and CR2 via a lentiviral vector. They found these treated cells continued to have disrupted CCR5 expression, did not exhibit mutations at highly homologous regions, and resisted HIV infection. Interestingly, they could not replicate these results in primary T cells, so there is still work to be done.
Description Size Cat. No.
Purified anti-CRISPR (CAS9) Antibody 100 µg 844301

CRISPR (CAS9) Data

(Left) HeLa cells expressing Tet-inducible Flag-tagged S. pyogenes Cas9 were seeded onto glass cover slips and induced with 1.5 µg/ml Doxycyclin for 30 hours. Cells were fixed, permeablized, and stained with anti-Cas9 (Clone 7A9) antibody. After incubation with donkey anti-mouse Alexa Fluor® 488, the nuclei were counter-stained with Hoechst 33342. (Right) HeLa cells were transiently transfected with Flag-tagged S. pyogenes Cas9 and lysed in hot Laemmli buffer for 30 hours post transfection. Equal amounts of untransfected and transfected cell lysates were separated by SDS-PAGE, transferred to a nitrocellulose membrane, and incubated with the indicated antibodies.

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