The genetic program of oocytes can be modified in vivo in the zebrafish ovary

• 来源期刊: 分子细胞生物学报（英文版）
• 年，卷（期）: 2018,（6）
• 页码范围: 479-493
• 页数: 15页
• 语种: 英文
• DOI: 10.1093/jmcb/mjy044

摘要

Oocytes,the irreplaceable gametes for generating a new organism,are matured in the ovary of living female animals.It is unknown whether any genetic manipulations can be applied to immature oocytes inside the living ovaries.As a proof-of-concept,we here demonstrate genetic amendments of zebrafish immature oocytes within the ovary.Oocyte microinjection in situ (OMIS) stimulates tissue repair responses,but some of the microinjected immature oocytes are matured,ovulated and fertilizable.By OMIS-mediated Cas9 approach,ntla and gata5 loci of oocytes arrested at prophase I of meiosis are successfully edited before fertilization.Through OMIS,high efficiency of biallelic mutations in single or multiple loci using Cas9/gRNAs allows immediate manifestation of mutant phenotypes in Fo embryos and multiple transgenes can co-express the reporters in Fo embryos with patterns similar to germline transgenic embryos.Furthermore,maternal knockdown of dnmt1 by antisense morpholino via OMIS results in a dramatic decrease of global DNA methylation level at the dome stage and causes embryonic lethalityprior to segmentation period.Therefore,OMIS opens a door to efficiently modify the genome and provides a possibility to repair genetically abnormal oocytes in situ.