Extended Data Fig. 9: Development of comodality experiment and QC analysis.

a, Proof-of-concept experiment to analyze SCITO-seq using ATAC-kit. UMAP of PBMCs stained with 12 representative surface markers (CD4, 8a, 14, 16, 45, 45RA, 45RO, 19, 20, 56, 11c, and HLA-DR) in 5 separate pools loading 5x10⁴ cells (nCM: non-conventional monocytes, cMono: conventional monocytes). B, Schematic of GEM ligation step in the comodality experiment using the 10x Genomics ATAC-seq kit. Detailed sequence structure of the RNA and ADT capture during the GEM reaction using the scifi-RNA-seq workflow. A more detailed workflow for the RNA can be found in the Supplementary Figure 2 in the scifi-RNA-seq paper. 10x_round2 refers to the 16 bp droplet barcode, round1 barcode refers to the well barcode (11 bp) used in the in situ reverse transcription reaction. Untemplated ‘CCC’ is added at the end of the reverse transcription reaction (MMLV variant). Antibody barcode (Ab BC 10-bp) and antibody handle (Ab handle 20-bp, conjugated directly to the blue antibody) sequences are specific to each antibody. Read2n represents Read2 Nextera sequence. Compared to the bridge oligo 1 (used to capture in-situ RT mRNA molecules), bridge oligo 2 has an extra 10-bp (AACGTATCGA between red and blue colored sequences). ddC (dideoxy C) and InvdT (inverted dT) for preventing extension. Arrow indicates the ligation site during the GEM reaction. c, Dimensional reduction using UMAP with normalized RNA counts and corresponding cell line specific ADT marker expressions on the UMAP space. d, Dimensional reduction using UMAP with normalized RNA counts and corresponding single RNA marker expressions on the UMAP space.