Table 1.

Summary of technical features of the scRNA-seq methods described in the review.

Methods	Summary	Advantages	Challenges
Smart-seq [10,11]	•102–103 cells/run •Detects full-length transcript •Addition of a few cytosines on 5′ end of full-length transcript allows hybridization with oligonucleotide primer	•Available commercial kits •Detection of different splice variants	•No detection of strand-specific nature of mRNAs
CEL-seq [12,13]	•102–103 cells/run •Only 3′-tag transcripts •Pipets single cell per tube	•Improved accuracy •Strand specificity and efficient barcoding	•Difficult to distinguish splice variants •Less sensitive
Qualtz-seq [14]	•104–105 cells/run •Cell isolation using FACS •Barcoding cells and first round of PCR performed on individual cell	•High UMI conversion efficiency •Low cell/run cost	•High amplification error rate •Smaller fragments preference
MARS-seq [15]	•103–5 × 103 cells/run •Cell isolation using FACS •Barcoding cells and first round of PCR performed on individual cell •Only 3′-tag transcripts	•Low reaction volume •Low noise •Strand specificity	•Not suitable for identifying splice variants •Limited to polyA RNAs •Requires FACS
Cyto-seq [16]	•102–104 cells/run •Only 3′-tag transcripts •PCR amplification using gene-specific primers •Beads with unique barcodes used for barcoding and transcript amplification	•High throughput •No restriction on cell sizes	•Time-consuming •Trade-off between sequencing depth and detection of differential gene expression
SUPeR-seq [17]	•~10 cells/run (micromanipulation) •Individual cell processing •Random primers with universal anchor sequence used for PCR amplification	•Detection of circular RNAs •3′ bias avoidable	•Low throughput
Drop-seq [18]	•Split and pool synthesis of cell barcodes and UMI synthesis conducted on primer beads •cDNA amplification of transcripts of the cells carried within droplets •Only 3′-tag transcripts	•Low cost •Robust cell processing (104 cells/day) •High yield •Customizable cell barcode	•High dependency on microfluidics
InDrop [19]	•Only 3′-tag transcripts •Polyacrylamide hydrogels with ssDNA primers with barcodes and polyT tails used •Each cell suspended in droplet with hydrogel and cell lysis proceeds within the droplet	•Low cell/run cost •Robust cell processing •High yield •Customizable cell barcode	•Low mRNA capture efficiency •One to one labeling of cell and barcode not guaranteed •High dependency on microfluidics
MATQ-seq [20]	•~102 cells/run •Cells mouth-pipetted into individual PCR tube •Barcodes incorporated to transcript from G enriched primers that bind to polyC tail	•Captures both polyA and non-polyA RNAs •Low 3′ end bias	•Low throughput
Chromium [21]	•102–104 cells/run •Only 3′-tag transcripts •Barcoded gel beads, cells and enzymes partitioned by oil	•Robust cell processing •Automated procedures •Relatively high cell capture efficiency	•High dependency on microfluidics
sci-RNA-seq [22]	•Methanol fixation of cells •Only 3′-tag transcripts •Reverse transcription incorporates UMI and barcode to each cell •Transposase used prior to library amplification	•Minimized perturbance to cell state or RNA integrity •FACS step can be incorporated	•Low throughput
Seq-Well [23]	•Method largely follows Drop-seq method •Cells loaded into subnano liter well by gravity	•Microfluidics device-independent •Potential for multi omics measurement at single cell scale	•Not fully automated
DroNC-seq [24]	•Method largely follows Drop-seq method •Only 3′-tag transcripts •New microfluidics design and nuclei isolation incorporated to the original Drop-seq method	•Reduced nuclei isolation time •Minimized RNA degradation	•High dependency on microfluidics
SPLiT-seq [25]	•~5 × 104 cells/run •Cell or nuclei are fixed with formaldehyde •Only 3′-tag transcripts •Transcriptome identification performed by four rounds of combinatorial barcoding •Barcoded samples undergo PCR amplification and are pooled to be sequenced	•Minimized perturbance to cell state or RNA integrity •Independent of microfluidics device	•Low number of average read/cell •Low cell type differentiation resolution