d Comparison of base editing outcomes at three pathogenic SNVs that can be created by five examined BEs in 293FT cells. c The diagram to illustrate preferentially targetable SNVs. The cytosines were counted with the base distal to the PAM setting as position 1 in Cas9-based BEs and with the base proximal to the PAM setting as position 1 in Cpf1-based BEs.
Mj human nature ly windows#
b The detailed target sites and editing windows of BE3, BE4max, eBE-S3, hA3A-eBE-Y130F, and dCpf1-eBE are shown. a The diagram to illustrate the use of BEs in creating or correcting C-to-T (G-to-A) SNVs or T-to-C (A-to-G) SNVs to model or correct disease-related mutations. We further profile the accessibilities of 20 BEs to all reported human pathogenic-related T-to-C or C-to-T point mutations in silico and build a BEable-GPS (Base Editable prediction of Global Pathogenic SNVs) database to provide a resource for potential gene therapies and biomedical studies.Ĭomparison of base editing outcomes at pathogenic SNVs. In this study, we experimentally compare a panel of five BEs for their editing efficiency and product purity at sites of human pathogenic C-to-T mutations that can be created or corrected by the same panel of BEs. More importantly, a database comprehensively cataloging pathogenic point mutations that can be corrected or created by different BEs has been lacking. However, BEs with different Cas proteins, e.g., Cas9 or Cas12a (also known as Cpf1), and different deaminases, e.g., rat APOBEC1 (rA1) or human APOBEC3A (hA3A), have not been directly compared for their utility in creating or correcting pathogenic point mutations. BEs hold the potential to be used for correcting and creating pathogenic point mutations (Fig. A uracil DNA glycosylase inhibitor (UGI) is fused to BEs to prevent unintended mutagenesis during the process of base editing, and additional UGIs co-expressed in trans with BEs (enhanced BE, eBE) further enhance the efficiency and fidelity of base editing.
Guided by the Cas moiety, BEs catalyze direct C-to-T changes with its fused cytidine deaminase moiety.
Distinct to Cas nucleases, which trigger homology-directed repair (HDR)-mediated gene correction by cleaving DNA double strands, BEs induce base changes in targeted genomic regions independent of the generation of DNA double-strand breaks (DSB) generally. A number of base editors (BEs), which combine different APOBEC (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like)/AID (activation-induced deaminase) cytidine deaminase family members with distinct CRISPR/Cas proteins, have been developed to achieve programmable C-to-T changes in different sequence contexts or backgrounds.
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