Single C-to-T substitution using engineered APOBEC3G-nCas9 base editors with minimum genome- and transcriptome-wide off-target effects

被引：69

作者：

Lee, Sangsin ^{[1
]}

Ding, Ning ^{[2
]}

Sun, Yidi ^{[3
,4
]}

Yuan, Tanglong ^{[5
]}

Li, Jing ^{[1
]}

Yuan, Qichen ^{[2
]}

Liu, Lizhong ^{[6
]}

Yang, Jie ^{[2
]}

Wang, Qian ^{[7
]}

Kolomeisky, Anatoly B. ^{[2
,7
,8
]}

Hilton, Isaac B. ^{[1
,6
]}

Zuo, Erwei ^{[5
]}

Gao, Xue ^{[1
,2
]}

机构：

[1] Rice Univ, Dept Bioengn, Houston, TX 77030 USA

[2] Rice Univ, Dept Chem & Biomol Engn, Houston, TX 77005 USA

[3] Univ Chinese Acad Sci, CAS Key Lab Syst Biol, CAS Ctr Excellence Mol Cell Sci, Chinese Acad Sci,Inst Biochem & Cell Biol,Shangha, Shanghai 200031, Peoples R China

[4] Univ Chinese Acad Sci, Chinese Acad Sci, Biomed Big Data Ctr,Shanghai Inst Nutr & Hlth,Sha, Key Lab Computat Biol,CAS MPG Partner Inst Comput, Shanghai 200031, Peoples R China

[5] Chinese Acad Agr Sci, Shenzhen Branch, Guangdong Lab Lingnan Modern Agr, Genome Anal Lab,Minist Agr,Agr Genom Inst Shenzhe, Shenzhen 518124, Peoples R China

[6] Rice Univ, Dept Biosci, Houston, TX 77005 USA

[7] Rice Univ, Ctr Theoret & Biol Phys, Houston, TX 77005 USA

[8] Rice Univ, Dept Chem, POB 1892, Houston, TX 77005 USA

来源：

SCIENCE ADVANCES | 2020年 / 6卷 / 29期

基金：

中国国家自然科学基金;

关键词：

DNA; DEAMINATION; DOMAIN;

D O I：

10.1126/sciadv.aba1773

中图分类号：

O [数理科学和化学]; P [天文学、地球科学]; Q [生物科学]; N [自然科学总论];

学科分类号：

07 ; 0710 ; 09 ;

摘要：

Cytosine base editors (CBEs) enable efficient cytidine-to-thymidine (C-to-T) substitutions at targeted loci without double-stranded breaks. However, current CBEs edit all Cs within their activity windows, generating undesired bystander mutations. In the most challenging circumstance, when a bystander C is adjacent to the targeted c, existing base editors fail to discriminate them and edit both Cs.To improve the precision of CBE, we identified and engineered the human APOBEC3G (A3G) deaminase; when fused to the Cas9 nickase, the resulting A3G-BEs exhibit selective editing of the second C in the 5'-CC-3' motif in human cells. Our A3G-BEs could install a single disease-associated C-to-T substitution with high precision. The percentage of perfectly modified alleles is more than 6000-fold for disease correction and more than 600-fold for disease modeling compared with BE4max. On the basis of the two-cell embryo injection method and RNA sequencing analysis, our A3G-BEs showed minimum genome- and transcriptome-wide off-target effects, achieving high targeting fidelity.

引用

页数：12

共 44 条

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