Fig. 2

Quality of single-indexed inDrop libraries sequenced alongside Illumina libraries and data loss from index hopping. a The base calling accuracy plot for a inDrop V2 library on a NextSeq sequencing run, depicting the spread of quality scores as each base is sequenced. This plot consists of a series of box plots where each box plot maps the percent of clusters in each image of the flow cell with quality scores ≥30 (called Q30) in each cycle. The first 100 cycles correspond to the transcript read; the next 6 correspond to the i7 index read; the final 50 correspond to the cell barcode + UMI reads. The last 6 cycles read into the poly A tail due to the variable length of the inDrop cell barcodes. b The base calling accuracy plot for a inDrop V2 library sequenced alongside the control Illumina library, PhiX, on a NextSeq. When sequencing alongside PhiX, the 7-base long i7- and i5- index reads are used so that PhiX reads can be filtered out and discarded during demultiplexing. c Plot of the calculated proportion of cell barcodes that need to be discarded from single-indexed sequencing runs at different levels of multiplexing. We assume each sample will contain ~ 3000 cell barcodes