INTACT was adapted from the method described in Pankova and Borst (2016) and pioneered in flies by Henry et al. (2012). The INTACT method of extracting nuclear RNA for sequencing provides several advantages compared to a general RNA-seq approach. First, the ability to use flash-frozen tissue, in contrast to FACS or dissected tissue samples, allows for an accurate examination of the current transcriptional profile of genetically targeted cells. Second, nuclear RNA from neurons contributes to the integrity of the active transcriptome snapshot by minimizing contamination from messenger RNA (mRNA) stored in the cytoplasm, along dendrites, and within axons, while allowing for the detection of experience-dependent differential expression (Lacar et al. 2016). Finally, given evidence that mRNA handling in subcellular compartments has been implicated in the formation and memory storage (Bramham and Wells 2007; Richter 2010; Shigeoka et al. 2016; Nakahata and Yasuda 2018; Biever et al. 2019), this approach provides a robust profile of the stable postexposure transcriptome unencumbered by the diversity of whole-cell RNA.
