| Products/Services Used | Details | Operation |
|---|---|---|
| Gene Synthesis | BE4max were obtained from Addgene (#112093). The DNA fragement of hA3G-CTD was synthesized and cloned into BE4max by Genscript Biotech (Nanjing) to create A3G-BE4max. The D316R/D317R, D316E or N244G mutations were induced into hA3G-CTD to produce A3G-D316R/D317R, A3G-D316E and A3G-N244G. Seven mutations (R1335A/L1111R/D1135V/G1218R/E1219F/A1322R/T1337R) of SpCas9 were introduced into BE4max and A3G-BE4max to create NG-BE4max and A3G-NG. Plasmid site-directed mutagenesis was performed using the Fast Site-Directed Mutagenesis Kit (TIANGEN, Beijing). All the site-directed mutation primers are listed in Table S3. The amino acid sequences of plasmids are listed in the Supplementary sequence. | Get A Quote |
Cytidine base editors, composed of a cytidine deaminase fused to Cas9 nickase, enable efficient C-to-T conversion in various organisms. However, current base editors can induce unwanted bystander C-to-T conversions when more than one C is present in the activity window of cytidine deaminase, which negatively affects the precision. Here, we develop a new base editor with CC context-specificity using rationally engineered human APOBEC3G, thus significantly reduce unwanted bystander activities. In addition, efficient C-to-T conversion that can further recognize relaxed NG PAMs is achieved by combining an engineered SpCas9-NG variant. These novel base editors with improved precision and targeting scope will expand ... More
