Engineering miniature IscB Nickase for Robust Base Editing with Broad Targeting Range
Robust miniature Cas-based transcriptional modulation by engineering Un1Cas12f1 and tethering Sso7d
Engineering Miniature CRISPR-Cas Un1Cas12f1 for Efficient Base Editing
Tracking paired-wise genomic loci by the ParB–ParS and Noc-NBS systems in living cells
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IminiCBE and SIminiCBE
2024.07.14
To improve the low editing efficiency of OgeuIscB (IscB from human gut metagenome) in mammalian cells, we developed high-efficiency miniature base editors by engineering OgeuIscB nickase and its cognate ωRNA, termed IminiBEs. And we fused non-specific DNA-binding protein Sso7d to IminiBEs increased the editing efficiency of low-efficiency sites by around two- to threefold, and we termed it SIminiBEs.IminiCBE and SIminiCBE recognize NNRR, NNRY and NNYR target-adjacent motifs, which broaden the canonical NWRRNA target-adjacent motif sites for the wild-type IscB nickase.
This work is published in NCB.
STUminiABE and STUminiCBE
2024.04.24
To deliver gene editing tools within single AAV, we developed hypercompact and high-efficiency base editors by engineering Un1Cas12f1, fusing non-specific DNA binding protein Sso7d, and truncating single guide RNA (sgRNA), termed STUminiBEs. We demonstrated robust A-to-G conversion (54% on average) by STUminiABEs or C-to-T conversion (45% on average) by STUminiCBEs. We packaged STUminiCBEs into AAVs and successfully introduced a premature stop codon on the PCSK9 gene in mammalian cells. In sum, we developed efficient miniature base editors and they could readily be packaged into AAVs for biological research or biomedical applications.
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SminiCRa and SminiCRi
2024.04.03
To deliver gene regulation tools within single aav and enable robust gene regulation, we developed a robust miniature Cas-based transcriptional activation or silencing system using Un1Cas12f1. Our tools reached a 5,628-fold activation of the ASCL1 gene and at least hundreds-fold activation at other genes examined.We generated an all-in-one AAV vector AIOminiCRi used to silence the disease-related gene SERPINA1. AIOminiCRi AAVs led to the 70% repression of the SERPINA1 gene in the Huh-7 cells. In summary, miniCRa, SminiCRa, miniCRi, and SminiCRi are robust miniature transcriptional modulators with high specificity that expand the toolbox for biomedical research and therapeutic applications.
For more detail, please read here