The zebrafish has become a popular vertebrate animal model in biomedical research.However,it is still challenging to make conditional gene knockout(CKO)models in zebrafish due to the low efficiency of homologous recom...The zebrafish has become a popular vertebrate animal model in biomedical research.However,it is still challenging to make conditional gene knockout(CKO)models in zebrafish due to the low efficiency of homologous recombination(HR).Here we report an efficient non-HR-based method for generating zebrafish carrying a CKO and knockin(KI)switch(zCKOIS)coupled with dual-color fluorescent reporters.Using this strategy,we generated hey2^zCKOIS which served as a hey2 KI reporter with EGFP expression.Upon Cre induction in targeted cells,the hey2^zCKOIS was switched to a non-functional CKO allele hey2^zCKOIS-inv associated with Tag RFP expression,enabling visualization of the CKO alleles.Thus,simplification of the design,and the visibility and combination of both CKO and KI alleles make our z CKOIS strategy an applicable CKO approach for zebrafish.展开更多
基金supported by the Young Scientists Fund of the National Natural Science Foundation of China(31500849)Shanghai Municipal Science and Technology Major Project(18JC1410100,2018SHZDZX05)+3 种基金the Key Research Program of Frontier Sciences(QYZDY-SSW-SMC028)the Strategic Priority Research Program(XDB32010200)of Chinese Academy of Sciencesthe International Partnership Program,Bureau of International Co-operation of Chinese Academy of Sciences(153D31KYSB20170059)China Wan-Ren Program,and Shanghai Leading Scientist Program
文摘The zebrafish has become a popular vertebrate animal model in biomedical research.However,it is still challenging to make conditional gene knockout(CKO)models in zebrafish due to the low efficiency of homologous recombination(HR).Here we report an efficient non-HR-based method for generating zebrafish carrying a CKO and knockin(KI)switch(zCKOIS)coupled with dual-color fluorescent reporters.Using this strategy,we generated hey2^zCKOIS which served as a hey2 KI reporter with EGFP expression.Upon Cre induction in targeted cells,the hey2^zCKOIS was switched to a non-functional CKO allele hey2^zCKOIS-inv associated with Tag RFP expression,enabling visualization of the CKO alleles.Thus,simplification of the design,and the visibility and combination of both CKO and KI alleles make our z CKOIS strategy an applicable CKO approach for zebrafish.
