Rationally engineered Staphylococcus aureus Cas9 nucleases with high genome-wide specificity.

RNA-guided CRISPR-Cas9 proteins have been widely used for genome editing, but their off-target activities limit broad application. The minimal Cas9 ortholog from Staphylococcus aureus (SaCas9) is commonly used for in vivo genome editing; however, no variant conferring high genome-wide specificity is available. Here, we report rationally engineered SaCas9 variants with highly specific genome-wide activity in human cells without compromising on-target efficiency. One engineered variant, referred to as SaCas9-HF, dramatically improved genome-wide targeting accuracy based on the genome-wide unbiased identification of double-stranded breaks enabled by sequencing (GUIDE-seq) method and targeted deep sequencing analyses. Among 15 tested human endogenous sites with the canonical NNGRRT protospacer adjacent motif (PAM), SaCas9-HF rendered no detectable off-target activities at 9 sites, minimal off-target activities at 6 sites, and comparable on-target efficiencies to those of wild-type SaCas9. Furthermore, among 4 known promiscuous targeting sites, SaCas9-HF profoundly reduced off-target activities compared with wild type. When delivered by an adeno-associated virus vector, SaCas9-HF also showed reduced off-target effects when targeting VEGFA in a human retinal pigmented epithelium cell line compared with wild type. Then, we further altered a previously described variant named KKH-SaCas9 that has a wider PAM recognition range. Similarly, the resulting KKH-HF remarkably reduced off-target activities and increased on- to off-target editing ratios. Our finding provides an alternative to wild-type SaCas9 for genome editing applications requiring exceptional genome-wide precision.

Author Correction: Gene and mutation independent therapy via CRISPR-Cas9 mediated cellular reprogramming in rod photoreceptors.

In the initial published version of this article, we inadvertently stated that "all procedures were conducted with the approval and under the supervision of the Institutional Animal Care and Use Committee (IACUC) at the University of California, San Diego". Given that all animal work that was conducted for this project was performed at the City University of Hong Kong and Guangzhou Women and Children's Medical Center, we would like to instead, acknowledge these programs for their oversight of the animal studies. This correction does not affect the description of the results or the conclusions of this work.

Acceptability of and willingness to pay for using a smartphone-based vaccination application in a Vietnamese cohort.

BACKGROUND: The increasing accessibility of smartphone in Vietnam shows potential in using smartphone applications for vaccination management to improve compliance. However, the acceptability and financial feasibility of using this application in Vietnam have not yet been understood. This study measured the general perception of and willingness to pay (WTP) for using smartphone-based vaccination management applications and their associated factors in Vietnam. SUBJECTS AND METHODS: A cross-sectional study was conducted between March and April 2016 in an urban vaccination clinic at the Hanoi Medical University in Vietnam. Convenience sampling was used to recruit 429 adult participants who had received vaccinations or whose children were vaccinated. Sociodemographic characteristics and the acceptability of and WTP for using smartphone-based vaccination management applications were evaluated. RESULTS: Among participants who used smartphone applications, 5% were aware of existing vaccination management applications, of whom 57.9% had previously used the applications. About 69.6% of participants believed that the applications were necessary, 93.7% of them were also willing to use the applications, and 79.1% were willing to pay 92.7 thousand Vietnamese dong (VND) for the applications on average. Participants who were older, unemployed, earned more household income, and having knowledge about functions and benefits of vaccination were less likely to use the vaccination applications. Participants who brought their children to get vaccinated at younger age and with higher level of education were willing to pay more for vaccination applications. CONCLUSION: High levels of acceptability of, willingness to use, and WTP for smartphone-based vaccination management applications among Vietnamese participants are reported. Increased education and awareness about the benefits of vaccination and vaccination management applications through community health workers might increase the feasibility of implementing such applications and perception toward their usage among young adult smartphone users. In addition, building a stronger relationship with health care providers at hospitals might improve the application's visibility and adoption.