| Autor: |
Zhang C; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China., Li J; Taishan College, Shandong University, Qingdao 266237, China., Tarique I; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China., Zhang Y; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China., Lu T; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China., Wang J; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China., Chen A; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China., Wen F; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China., Zhang Z; Taishan College, Shandong University, Qingdao 266237, China., Zhang Y; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China., Shao M; Shandong Provincial Key Laboratory of Animal Cell and Developmental Biology and Key Laboratory for Experimental Teratology of the Ministry of Education, School of Life Sciences, Shandong University, Qingdao 266237, China.; Taishan College, Shandong University, Qingdao 266237, China. |
| Abstrakt: |
Maternal products are those mRNAs and proteins deposited during oogenesis, which play critical roles in controlling oocyte formation, fertilization, and early embryonic development. However, loss-of-function studies for these maternal factors are still lacking, mainly because of the prolonged period of transgenerational screening and technical barriers that prevent the generation of maternal (M) and maternal and zygotic (MZ) mutant embryos. By the transgenic expression of multiple sgRNAs targeting a single gene of interest in the background of a transgenic line Tg( zpc : zcas9 ) with oocyte-specific cas9 expression, we have successfully obtained maternal or maternal-zygotic mutant for single genes in F1 embryos. In this work, we tandemly connected a maternal GFP marker and eight sgRNA expression units to target dvl2 and dvl3a simultaneously and introduced this construct to the genome of Tg( zpc : zcas9 ) by meganuclease I- Sce I. As expected, we confirmed the existence of M dvl2 ;M dvl3a embryos with strong defective convergence and extension movement during gastrulation among outcrossed GFP positive F1 offspring. The MZ dvl2 ;MZ dvl3a embryos were also obtained by crossing the mutant carrying mosaic F0 female with dvl2 +/- ; dvl3a -/- male fish. This proof-of-principle thus highlights the potential of this conditional knockout strategy to circumvent the current difficulty in the study of genes with multiple functionally redundant paralogs. |
