Correction of the pathogenic mutation in the G6PC3 gene by adenine base editing in mutant embryos.
10.3760/cma.j.issn.0253-2727.2023.04.008
- Author:
Man HONG
1
;
Ping WANG
2
;
Tao SHANGGUAN
3
;
Guang Lei LI
4
;
Rui Peng BIAN
1
;
Wei HE
3
;
Wen JIANG
2
;
Jie Ping CHEN
2
Author Information
1. Guizhou University Medical College, Guiyang 550025, China.
2. Department of Hematology, the First Affiliated Hospital of Army Medical University (Southwest Hospital), Chongqing 400038, China.
3. Department of Reproductive Medicine, the First Affiliated Hospital of Army Medical University (Southwest Hospital), Chongqing 400038, China.
4. Department of Reproductive Medicine, Third Affiliated Hospital of Guangzhou Medical University, Guangzhou 510150, China.
- Publication Type:Journal Article
- Keywords:
Base editing;
Embryo correction;
Gene, G6PC3;
Severe congenital neutropenia
- MeSH:
Humans;
Gene Editing;
CRISPR-Cas Systems;
Adenine;
HEK293 Cells;
Mutation;
Glucose-6-Phosphatase/metabolism*
- From:
Chinese Journal of Hematology
2023;44(4):308-315
- CountryChina
- Language:Chinese
-
Abstract:
Objective: To determine whether the adenine base editor (ABE7.10) can be used to fix harmful mutations in the human G6PC3 gene. Methods: To investigate the safety of base-edited embryos, off-target analysis by deep sequencing was used to examine the feasibility and editing efficiency of various sgRNA expression vectors. The human HEK293T mutation models and human embryos were also used to test the feasibility and editing efficiency of correction. Results: ①The G6PC3(C295T) mutant cell model was successfully created. ②In the G6PC3(C295T) mutant cell model, three distinct Re-sgRNAs were created and corrected, with base correction efficiency ranging from 8.79% to 19.56% . ③ ABE7.10 could successfully fix mutant bases in the human pathogenic embryo test; however, base editing events had also happened in other locations. ④ With the exception of one noncoding site, which had a high safety rate, deep sequencing analysis revealed that the detection of 32 probable off-target sites was <0.5% . Conclusion: This study proposes a new base correction strategy based on human pathogenic embryos; however, it also produces a certain nontarget site editing, which needs to be further analyzed on the PAM site or editor window.
