Base editing, as an expanded clustered regularly interspaced short palindromic repeats(CRISPR)-Cas genome editing strategy, permits precise and irreversible nucleotide conversion. SaKKH, an efficient variant of a Cas9...Base editing, as an expanded clustered regularly interspaced short palindromic repeats(CRISPR)-Cas genome editing strategy, permits precise and irreversible nucleotide conversion. SaKKH, an efficient variant of a Cas9 ortholog from Staphylococcus aureus(SaCas9), is important in genome editing because it can edit sites with HHHAAT protospacer adjacent motif(PAM) that the canonical Streptococcus pyogenes Cas9(SpCas9) or its variants(e.g. xCas9, Cas9-NG) cannot. However, several technical parameters of SaKKH involved base editors have not been well defined and this uncertainty limits their application. We developed an effective multiplex cytosine base editor(SaKKHn-pBE) and showed that it recognized NNARRT, NNCRRT, NNGRGT, and NNTRGT PAMs. Based on 27 targets tested, we defined technical parameters of SaKKHn-pBE including the editing window, the preferred sequence context, and the mutation type. The editing efficiency was further improved by modification of the SaKKH sgRNA. These advances can be applied in future research and molecular breeding in rice and other plants.展开更多
Dear Editor,Genome editing tools are leading a revolution in plant breeding.In particular,prime editors(PEs)can install all types of base changes and small insertions/deletions at precise positions in plant genomes(An...Dear Editor,Genome editing tools are leading a revolution in plant breeding.In particular,prime editors(PEs)can install all types of base changes and small insertions/deletions at precise positions in plant genomes(Anzalone et al.,2019).PEs are by far the most powerful approach for improving traits conferred by gain-of-function point mutations.Early versions of PEs suffered from low editing efficiency,but the latest PEs can perform edits at a much higher efficiency thanks to the extensive efforts of re-searchers from around the world.Most modifications to improve PE efficiency have focused on the optimization of PE protein components and structure.展开更多
基金supported by the Beijing Scholars Program[BSP041]。
文摘Base editing, as an expanded clustered regularly interspaced short palindromic repeats(CRISPR)-Cas genome editing strategy, permits precise and irreversible nucleotide conversion. SaKKH, an efficient variant of a Cas9 ortholog from Staphylococcus aureus(SaCas9), is important in genome editing because it can edit sites with HHHAAT protospacer adjacent motif(PAM) that the canonical Streptococcus pyogenes Cas9(SpCas9) or its variants(e.g. xCas9, Cas9-NG) cannot. However, several technical parameters of SaKKH involved base editors have not been well defined and this uncertainty limits their application. We developed an effective multiplex cytosine base editor(SaKKHn-pBE) and showed that it recognized NNARRT, NNCRRT, NNGRGT, and NNTRGT PAMs. Based on 27 targets tested, we defined technical parameters of SaKKHn-pBE including the editing window, the preferred sequence context, and the mutation type. The editing efficiency was further improved by modification of the SaKKH sgRNA. These advances can be applied in future research and molecular breeding in rice and other plants.
基金supported by grants from the National Key Research and Development Program of China(2023YFD1202905)the National Natural Science Foundation of China(32272629).
文摘Dear Editor,Genome editing tools are leading a revolution in plant breeding.In particular,prime editors(PEs)can install all types of base changes and small insertions/deletions at precise positions in plant genomes(Anzalone et al.,2019).PEs are by far the most powerful approach for improving traits conferred by gain-of-function point mutations.Early versions of PEs suffered from low editing efficiency,but the latest PEs can perform edits at a much higher efficiency thanks to the extensive efforts of re-searchers from around the world.Most modifications to improve PE efficiency have focused on the optimization of PE protein components and structure.
