Comment[GEAAccession]	E-GEAD-470
MAGE-TAB Version	1.1
Investigation Title	scRNAseq from mouse paraventricular nucleus of thalamus
Experiment Description	scRNAseq data from mouse paraventricular nucleus of thalamus (PVT). PVT from 6 (3 males and 3 females) were isolated from brain slices and tritulated to single cells. Cells were sorted to 384 well plate with FACS and sequencing libraries were constructed with Quartz-seq2 mehtod.
Experimental Design	cell type comparison design	translational bias design	replicate design
Experimental Factor Name	isolate
Experimental Factor Type	isolate
Person Last Name	Shima
Person First Name	Yasuyuki
Person Affiliation	Neurodegenerative Disorders Collaboration Laboratory  Center for Brain Science, RIKEN
Person Roles	submitter
Public Release Date	2022-04-26
Protocol Name	P-GEAD-1034	P-GEAD-1035	P-GEAD-1036	P-GEAD-1037	P-GEAD-1038
Protocol Type	sample collection protocol	nucleic acid extraction protocol	nucleic acid library construction protocol	nucleic acid sequencing protocol	normalization data transformation protocol
Protocol Description	We used 6 mice (3 males and 3 females of Calb2-Cre/+; Ai3/+, C57Bl6/J background, age: 118-151 days old) for scRNAseq. After slicing brains, we followed the single-cell preparation protocol previously described (Hempel et al., 2007). Slices were incubated in bubbled HEPES-ACSF with 1 mg/ml pronase for 70 minutes at room temperature. PVT was isolated with a scalpel under a fluorescent dissection microscope.  The isolated tissues were transferred to 1 % FBS/HEPES-ACSF and triturated with a series of fire-polished Pasteur pipettes with different tip diameters.FACS-ARIA II (Beckman Coulter) fluorescent cell sorter was used to sort live single cells (Hoechst +, 7-AAD -) to 384-well plates.	We followed a protocol described in Sasagawa et al. (doi: 10.1186/s13059-018-1407-3). Cell lysis and reverse transcription took place in each 384 well.	We followed a protocol described in Sasagawa et al. (doi: 10.1186/s13059-018-1407-3). cDNA were pooled into a single tube and following reactions (tagging, whole transcriptome amplificaiton, sonication, and ligation/PCR) were performed.	The indexed libraries were sequenced with NextSeq 500 (Illumina). Sequencing cycles were performed as following: Read1, 23 cycles; Index1, 6 cycles; Read2, 63 cycles.	The Quartz-seq2 pipeline (https://github.com/rikenbit/Q2-pipeline) was used to generate a digital expression matrix from BCL files. For transcript mapping with RNA-star (Dobin et al., 2013), we used a custom index file made from a reference mouse genome (mm10/GRCm38) and transgenes (EGFP and Cre).
SDRF File	E-GEAD-470.sdrf.txt
Comment[AEExperimentType]	RNA-seq of coding RNA from single cells
Comment[BioProject]	PRJDB12694
Comment[Last Update Date]	2022-04-26