| First Author | Rumpler É | Year | 2023 |
| Journal | J Biol Chem | Volume | 299 |
| Issue | 9 | Pages | 105121 |
| PubMed ID | 37536628 | Mgi Jnum | J:352799 |
| Mgi Id | MGI:7527776 | Doi | 10.1016/j.jbc.2023.105121 |
| Citation | Rumpler E, et al. (2023) Development of a versatile LCM-Seq method for spatial transcriptomics of fluorescently tagged cholinergic neuron populations. J Biol Chem 299(9):105121 |
| abstractText | Single-cell transcriptomics are powerful tools to define neuronal cell types based on co-expressed gene clusters. Limited RNA input in these technologies necessarily compromises transcriptome coverage and accuracy of differential expression analysis. We propose that bulk RNA-Seq of neuronal pools defined by spatial position offers an alternative strategy to overcome these technical limitations. We report a laser-capture microdissection (LCM)-Seq method which allows deep transcriptome profiling of fluorescently tagged neuron populations isolated with LCM from histological sections of transgenic mice. Mild formaldehyde fixation of ZsGreen marker protein, LCM sampling of approximately 300 pooled neurons, followed by RNA isolation, library preparation and RNA-Seq with methods optimized for nanogram amounts of moderately degraded RNA enabled us to detect approximately 15,000 different transcripts in fluorescently labeled cholinergic neuron populations. The LCM-Seq approach showed excellent accuracy in quantitative studies, allowing us to detect 2891 transcripts expressed differentially between the spatially defined and clinically relevant cholinergic neuron populations of the dorsal caudate-putamen and medial septum. In summary, the LCM-Seq method we report in this study is a versatile, sensitive, and accurate bulk sequencing approach to study the transcriptome profile and differential gene expression of fluorescently tagged neuronal populations isolated from transgenic mice with high spatial precision. |
