Delivery of CRISPR (clustered regularly interspaced short palindromic repeats)/CRISPR-associated protein-9 (Cas9) represents a major hurdle for successful clinical translation of genome editing tools. Owing to the large size of plasmids that encode Cas9 and single-guide RNA (sgRNA), genome editing efficiency mediated by current delivery carriers is still unsatisfactory to meet the requirement for its real applications. Herein, cationic polymer polyethyleneimine-β-cyclodextrin (PC), known to be efficient for small plasmid transfection, is reported to likewise mediate efficient delivery of plasmid encoding Cas9 and sgRNA. Whereas PC can condense and encapsulate large plasmids at high N/P ratio, the delivery of plasmid results in efficient editing at two genome loci, namely, hemoglobin subunit beta (19.1%) and rhomboid 5 homolog 1 (RHBDF1) (7.0%). Sanger sequencing further confirms the successful genome editing at these loci. This study defines a new strategy for the delivery of the large plasmid encoding Cas9/sgRNA for efficient genome editing.

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The adenosine base editor (ABE) is able to catalyze A•T to C•G conversion efficiently and precisely in vivo, representing a new method for gene therapy. Adeno associated virus (AAV) is a well‐studied vector for gene delivery in vivo. However, due to the limited loading capacity of AAV vector (≈4800 bp), it is difficult to package ABE (≈5400 bp) into a single AAV. To tackle this problem, ABE can be split into two smaller parts through intein‐mediated protein trans‐splicing. Here, 14 different split sites of nCas9 (Cas9 nickase) in combination with three different inteins (Mxe, Npu, and Rma) are screened through a GFP‐based reporter system to identify novel split‐ABEs. After infecting HEK293T and HeLa cells with dual AAVs, two split‐ABEs (split‐ABE‐Rma573 and split‐ABE‐Rma674) that can edit the target gene efficiently are identified. Furthermore, these dual‐AAV split‐ABEs can effectively disrupt the splicing acceptor of PCSK9 in mouse liver and the splicing donor of NR2E3 in mouse retina through AI‐MAST strategy. This study provides two new split‐ABEs to investigate gene function in vivo and in gene therapy, representing a new method to treat diseases by precisely repairing point mutations or inactivating genes through the AI‐MAST strategy.

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Oxygen vacancies in transition-metal oxides can facilitate photocatalysis activity critical for environmental remediation and solar energy conversion. Investigation of photocatalytic activity of TiO 2 with different amounts of the oxygen vacancies can provide fundamental insights into the oxygen-vacancy-dependent reaction mechanisms. Here, the oxygen vacancies in TiO 2 nanoparticles were investigated by the combination of transmission electron microscopy with electron energy loss spectra, x-ray near-edge structures, and x-ray absorption techniques. The results demonstrate that the oxygen vacancies mainly exist at the surface of TiO 2 and the crystalline structure of TiO 2 almost keeps no change during annealing under the different oxygen pressures at 500 • C. The exact quantity and distribution of the oxygen vacancies in TiO 2 nanoparticles were obtained. The surface oxygen vacancies on TiO 2 significantly enhance photocatalytic activity for Rhodamine B degradation and hydrogen generation. The formation of oxygen vacancies at the TiO 2 surfaces is responsible for engineering the band gap of TiO 2 and tailoring their electronic structures, and promoting light absorption and enhancement photocatalysis. This work could provide new insights into the understanding of the photocatalytic activity enhancement by the surface oxygen vacancies.