Journal: bioRxiv

Article Title: MULTI-seq: Scalable sample multiplexing for single-cell RNA sequencing using lipid-tagged indices

doi: 10.1101/387241

Figure Lengend Snippet: (A) Schematic overview of PDX experiment. Primary tumors and lung tissue from PDX mouse models were dissected and cryopreserved until the day of the experiment. These tissues were then thawed and dissociated prior to labeling with viability dyes, species-specific antibodies, and sample-specific MULTI-seq barcodes. Live hCD298+ and mCD45+ cells were then FACS-enriched and pooled prior to sequencing. (B) MULTI-seq sample classifications mapped onto barcode space. (C) Species and mouse/human doublet classifications from transcriptome data mapped onto barcode space. (D) Bar plots describing the proportion of mouse (tan) and human (green) cells loaded into the droplet microfluidic device (IN) compared to the species proportions in the final dataset (OUT).

Article Snippet: While existing methods are optimally configured to assay many thousands of cells, library preparation practices and the physical constraints of current commercially-available microfluidic devices (e.g., the Fluidigm C1 and 10X Genomics Single-Cell V2 systems) limit analyses to sets of 8 or fewer conditions in a typical scRNA-seq experiment.

Techniques: Labeling, Sequencing

Journal: Nucleic Acids Research

Article Title: A cost effective 5΄ selective single cell transcriptome profiling approach with improved UMI design

doi: 10.1093/nar/gkw1242

Figure Lengend Snippet: On chip barcoding workflow. After cell lysis in 4.5 nl poly-adenylated RNA is reverse-transcribed in 31.5 nl with an anchored oligodT primer. A PCR primer sequence and unique molecular identifiers (UMIs) are added to the 3΄ end of the cDNA via reverse transcriptase template switching. The cDNA is subsequently amplified and cell index sequences (barcode) as well as terminal biotins are introduced by PCR in the microfluidic device. The barcoded cDNAs are pooled, fragmented by tagmentation with Tn5 transposase and the biotinylated terminal fragments are isolated on streptavidin beads. 5΄ terminal fragments are selectively amplified and additional sequences required for Ion Torrent sequencers are introduced by PCR. For a detailed protocol see and for Illumina sequencers see .

Article Snippet: We sought to design a 5 ́ selective library preparation strategy that uses UMIs and fulfills the following criteria: (i) unbiased introduction of cell indices before the costly and labor intensive fragmentation step; (ii) compatibility with the Fluidigm C1 microfluidic device design; (iii) essentially sequencing platform independent; (iv) cost and labor effective.

Techniques: Lysis, Reverse Transcription, Sequencing, Amplification, Isolation