title: GSM8337513: Knockout; Mus musculus; RNA-Seq
library_construction_protocol: E10.5 embryos were dissected in cold PBS and the yolk sac was removed for genotyping. The embryos were stored in 1% FBS (Cytvia, SH30910.03) on ice for 3 hours during genotyping. The yolk DNA was extracted and processed for genotyping with QuickExtract DNA Extraction Solution (Epicentre, QE09050). Embryonic hearts from embryos with desired genotypes were further microdissected. Four embryonic hearts were pooled together for each group. The dissected tissue was incubated in 200 L TrypLE (ThermoFisher Scientific, 12563029) for 5 minutes. The samples were triturated with a 200 L pipette tip and incubated for 5 minutes. After the incubation period, 600 L of PBS/1% FBS was added, and the cells were strained through a 70 m cell strainer (BD Falcon, 352350) and subsequently centrifuged at 150 x g for 3 minutes, supernatant removed and pelleted cells resuspended in 35 L PBS/1% FBS as described previously (24). Cell viability was assessed with Cellometer Auto T4 (Nexcelom bioscience). Samples were used to prepare single cell droplet libraries in the 10X Genomics Chromium controller according to the Chromium Single Cell 3' Reagent Kit User Guide with reagents from Single Cell 3' Reagent Kit v3.1 (201456), library construction kit (201781), dual index plate TT set A (163910), and Chromium next gem chip G single cell kit (201862). Libraries were sequenced on the Illumina NextSeq 2000 and P2 100 cycle kit. Libraries were sequenced to 100 bp read length and 500 million read number.