By leveraging Takara Bio's expertise in single-cell RNA-seq library construction and the cell isolation and selection capabilities of the SMARTer ICELL8 systems, the SMARTer ICELL8 3' DE workflows enable high-throughput preparation of Illumina libraries for differential expression analysis of single cells from up to eight different samples in parallel.
The SMARTer ICELL8 Chip and Reagent 3' DE Kit (Cat. # 640164, 640165, or 640166) is compatible with the SMARTer ICELL8 Single-Cell System (purchased separately). Included in the kit is a SMARTer ICELL8 3' DE Chip, a 5,184-nanowell SmartChip that contains preprinted oligo(dT) primers, and reagents for cDNA synthesis and Illumina library prep.
Following staining and dilution, sample cells are dispensed into chip nanowells using the ICELL8 MultiSample NanoDispenser (MSND), and wells containing single, viable cells are selected using CellSelect Software provided with the SMARTer ICELL8 system. Following selection of cells for processing and analysis, RT-PCR reagents are dispensed into the corresponding wells, and cDNA synthesis and amplification are performed in-chip in a single run. Following RT-PCR, cDNA products are pooled and purified, and sequencing libraries are prepared using a transposase-based method that enriches for cDNA derived from mRNA 3' ends, followed by a limited-cycle PCR amplification step that incorporates Illumina index sequences.
The SMARTer ICELL8 cx 3' DE workflow for the SMARTer ICELL8 cx Single-Cell System (purchased separately; coming soon) requires the ICELL8 Reagent 3' DE Kit (Cat. # 640167), the SMARTer ICELL8 cx 3' DE Chip (Cat. # 640199), the SMARTer ICELL8 cx Loading Kit (Cat. # 640197), and the SMARTer ICELL8 Collection Kit (Cat. # 640048)—each purchased separately. The SMARTer ICELL8 cx Single-Cell System offers the same powerful technology for single-cell analysis as the system described above, but in an integrated automation platform which combines imaging and dispensing in one unit.
Sequencing data from the resulting libraries produced by either system can be traced back to individual cells using the nanowell-specific barcode sequences and can be used to perform differential gene expression analysis by mRNA 3' end-counting. By yielding high-quality, 3'-focused sequencing data from hundreds of cells in parallel, these kits provide an efficient, cost-effective solution for high-throughput differential expression analysis of single cells.