Combination of Single cell RNA-seq & Electrophysiological recording (PMID: 26689544)

Paper: Integration of electrophysiological recordings with single-cell RNA-seq data identifies neuronal subtypes. Nature biotechnology. (2016)
Organism: Mus Musculus
Cell types: Aspirates from cortical neurons
Sample number: 83
Library preparation: STRT-seq
DATA: GSE70844

  1. Patchs-seq = Pache-clamp stimulus protocol followed by the aspiration of the entire somatic compartment into the recording pipette
  2. CCK(BAC-dsRed)xGAD67(KI-GFP+/-) mouse
  3. Use patch-clamp for classify interneurons in cortical L1 into 5 subclasses
  4. Continuum of positive voltage pulses reduce the loss of RNA, which is negatively charged
  5. 1.6M reads/cell, 40% is uniquely mapped to 2,068 genes, around 6,000 mRNA molecules/cell
  6. Efficiency of RNA capturing is 7% (20% efficiency / 19,000 mRNA molecules/ 5,000genes with normal scSTRT)
  7. Discard cells with <= 1500 mRNA molecules without mt/repeated/rRNA
  8. Assign each neuron from Patch-seq to one of possible neuronal subtypes in previous paper PMID:25700174 by correlation to increase reliability
  9. i) separate to 2 groups, S1PyrL1-L6 or Int1-16, ii) classify by correlation, iii) groups for comparison are continuously updated (consist of 7 detail steps in all),
  10. Correlation between gene expression and electrophysiological parameters show significance (24 genes show significant correlation with electrophysiology parameter)
  11. Detect subtype-specific receptors difference
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