Targeted In Utero Electroporation of Somatosensory Cortex

IUE targeting the somatosensory cortex was performed using our previously published methods with minor modifications^{20 ,21 (link)}. Pregnant C57/BL6 mice were anesthetized at embryonic day 15 (E15) by intraperitoneal administration of a mixed solution of Ketamine HCl (100 mg/kg), Xylazine HCl (7.5 mg/kg), and Buprenorphine HCl (0.05 mg/kg). After the uterine horn was exposed by laparotomy, the shRNA plasmid (1 µg/µl) together with CAG promoter-driven GFP expression plasmids (1 µg/µl) (molar ratio, approximately 1:1) were injected into the lateral ventricles with a glass micropipette made from a micropillary tube (Narishige, Cat #GD-1), and electroporated into the VZ of the CP at E15. C11orf46 expression constructs (1 µg/µl) were also co-introduced for rescue experiments. For in vivo epigenomic editing, CAG-dCas9-ST (1 µg/µl), scFv-C11orf46 (1 µg/µl) and sgRNAs (1 µg/µl) together with shRNAs and GFP expression plasmid will be introduced at E15. For overexpression of Sema6a, CAG-Sema6a (2 µg/µl) or tdTomato (1 µg/µl) with GFP expression plasmid will be introduced at E15. For electroporation, Electrode pulses (40 V; 50 ms) were charged four times at intervals of 950 ms with an electroporator (Nepagene, Cat #CUY21EDIT). All experiments were performed in accordance with the institutional guidelines for animal experiments of Johns Hopkins University.