Direct gene disruption by TALENs in medaka embryos

作者：Wang Tiansu; Hong Yunhan^*

来源：Gene, 2014, 543(1): 28-33.

DOI：10.1016/j.gene.2014.04.013

• dnd
• Gene disruption
• Genome editing
• Gene targeting
• TALEN

Targeted gene disruption (GD) is powerful for generating genetic alterations in animal genomes. Engineered endonucleases such as zinc finger nucleases and transcription activator-like effector nucleases (TALENs) allow for GD directly in animal embryos to achieve germline transmission. Here we report procedures and parameters of TALEN-mediated GD in the fish medaka by using a germ cell-specific gene dnd as a model. Embryos at the 1-cell stage were microinjected with synthetic TALEN mRNAs and examined for the survival rate and GD efficiency. Medaka embryos can tolerate a high dosage of TALEN-mRNA injection and exhibit a steadily increasing GD efficiency with increasing mRNA dosages before peaking at 100 ng/mu l. This dosage produced similar to 24% efficiency for somatic GD. Some of the animals from manipulated embryos developed into fertile female and male. Most importantly, four fish (3 males and 1 female) examined by progeny-test were able to produce GD-bearing male and female gametes for germline transmission to F-1 generation at similar to 10% efficiency. Therefore, TALEN is proficient for somatic and germline GD in medaka embryos, and disruption of one dnd copy does not compromise somatic development and gamete production.

• 出版日期2014-6-10