Highly efficient RNA-guided base editing in rabbit

Abstract

Cytidine base editors (CBEs) and adenine base editors (ABEs), composed of a cytidine deaminase or an evolved adenine deaminase fused to Cas9 nickase, enable the conversion of C·G to T·A or A·T to G·C base pair in organisms, respectively. Here, we show that BE3 and ABE7.10 systems can achieve a targeted mutation efficiency of 53–88% and 44–100%, respectively, in both blastocysts and Founder (F0) rabbits. Meanwhile, this strategy can be used to precisely mimic human pathologies by efficiently inducing nonsense or missense mutations as well as RNA mis-splicing in rabbit. In addition, the reduced frequencies of indels with higher product purity are also determined in rabbit blastocysts by BE4-Gam, which is an updated version of the BE3 system. Collectively, this work provides a simple and efficient method for targeted point mutations and generation of disease models in rabbit.

Expanding the base editing scope in rice by using Cas9 variants

“…Therefore, the base editing windows for ABEs and CBEs at some target sites in our study (Table ) are larger than previously defined in mammalian systems (Gaudelli et al ., ; Komor et al ., ). Two recent in vivo base editing studies in mouse and rabbit also showed that ABEs and CBEs with SpCas9 can edit target adenines and cytosines outside of the canonical base editing activity windows at some target sites (Liu et al ., ,b). As the base editing windows characterized in previous studies were derived from limited target sites and were mainly from cell‐based assays which had relatively a short time for base editors to function (Gaudelli et al ., ; Komor et al ., ), more target sites need to be tested, and in vivo base editing studies are required to accurately define the base editing windows for both adenine and cytosine base editors.…”

Expanding targeting scope, editing window, and base transition capability of base editing in Corynebacterium glutamicum

“…However, for the majority of targets, the editing efficiencies were lower than 20%, which were not as high as those obtained in eukaryotes, such as approximately 50% in human cells (Gaudelli et al, 2017) and 44 to 100% in rabbits (Z. Liu et al, 2018). In nature, adenosine deaminase mediates deamination of adenosine (A) in RNA into inosine (I).…”

Modeling human point mutation diseases inXenopus tropicaliswith a modified CRISPR/Cas9 system

“…The effectiveness of these tools in X. tropicalis remains to be tested. 26 (2) numbers are shown on the left side. Genotype frequencies are shown on the right side.…”

“…Recently, base editor 3 (BE3), a fusion of a clustered regularly interspaced short palindromic repeat-associated protein 9 (Cas9) nickase, a cytidine deaminase [rat apolipoprotein B mRNA editing enzyme catalytic subunit 1 (APOBEC1)], and the uracil DNA glycosylase inhibitor (UGI), has been developed as a base editor (BE3), which efficiently induces the C-G pair to T-A pair transition in living human cells without inducing double-stranded DNA breaks (17). Subsequent studies demonstrated the effectiveness of this base editing strategy in yeasts, plants, and various animal models (18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28). Its application in X. tropicalis remains to be tested.…”