NET-Seq
Div-Seq
Div-Seq is a single-nucleus RNA sequencing technique that improves upon Nuc-Seq by incorporating 5-ethynyl-2’-deoxyuridine (EdU) labeling to identify dividing cells during their different cell stages. EdU labeling also enables identification of different cell types in complex tissue samples and rare cell populationsduring FACS. Briefly, samples are labeled in vivo with EdU, dissected, and fixed before isolation into single nuclei. Individual nuclei are tagged fluorescently and sorted by FACS. From this step, the procedure follows the Nuc-Seq method: cDNA synthesis is performed using the Smart-seq2 protocol, while the cDNA library is prepared with Tn5 transposase–mediated tagmentation.
Similar methods: Nuc-seq, snRNA-seq
Pros:
- Tracks transcriptome dynamics in single nuclei
- Detects rare cell populations
- Compatible with fresh, frozen, or fixed sample types
- EdU labeling gives unbiased identification of different types of dividing cells and their current stage in the cell cycle
- Mild nuclear dissociation technique minimizes gene expression changes commonly seen in protease-mediated dissociation
Cons:
- Excludes any information from cytoplasmic RNA