GSM6504521: Ascl1-mutAscl1-Myod1, 72h scRNA-seq; Mus musculus; RNA-Seq
Library preparation was performed according to the manufacter's instructions (Single Cell Multiome ATAC + Gene Expression, 10x Genomics). Briefly, nuclei were isolated from MEFs, resuspended in a transposition master mix and incubated at 60°C for 60 min. Transposed nuclei were added to the master mix and loaded together with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The poly-A RNA from the cell lysate contained in every single GEM was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The transposed genomic DNA was barcoded during transposition and contains Illumina R1, R2 and P5 primer sequences as well as  a 10x Barcode. Both transposed DNA and retrotranscribed cDNA were pre-amplified via PCR. ATAC-seq libraries were generated by a final PCR during which a P7 primer sequence and an i7 sample index are added. Gene expression libraries were generated through further amplification of the cDNA by PCR followed by enzymatic fragmentation and final PCR amplification adding both P5 and P7 Illumina primer sequences as well as i5 and i7 indexes. Library quality was validated using the Agilent High Sensitivity DNA Kit.