GSM9177145: mouse frontal cortex, RNA, biology_rep1; Mus musculus; OTHER
The frontal cortex were dissected by tweezer from adult mouse, then were treated by douncer to get the cells. Cells were crosslinked by formaldehyde and quenched by Tris glycine. Nuclei were extracted by NPB buffer from the cells. Nuclei were tagmentated by barcoded Tn5, RNA in the nuclei were reverse transcribed by barcoded primers. By doing two rounds of pool split and ligation, ligated the barcoded adaptor2 and adaptor3 to Tn5 adaptors and reverse transcription primers. Then did another round of pool split, each well contained 5000-90000 nuclei. Treated nuclei by SDS to release hybrid cDNA to supernatant from the nuclei. For the cDNA part, we performed reverse crosslink, template switch, double strand cDNA amplification, Tagmentation, PCR adding the 4th barcode for RNA library construction similar with split-seq. For the nuclei, we did in-situ digestion, in situ ligation, reverse crosslink, NlaIII digestion, circularization, PCR adding 4th barcode for DNA library construction similar with bulk HiCAR.