{"database": "metadata", "table": "run_metadata", "rows": [[76822, "SRR25374468", "SRX21113430", "SRS18382024", "SRP450614", "PRJNA996961", "Single cell transcriptome profiling of zebrafish pancreatic beta cells segregated by CaMPARI based calcium labeling.", "GSE237867", "Transcriptome Analysis", "Coordination of cell activity trough Ca2+ waves enables pancreatic \u00df cells to secrete precise quantities of insulin in response to blood sugar. However  how the coordinating Ca2+ waves are orchestrated in space and time remains unknown. By applying functional Ca2+ imaging  using CaMPARI and single molecule RNA detection  we reveal a gene signature depicting a molecularly and metabolically distinct state of first responder \u00df cells.  We highlight Vitamin B6 metabolism as a signature pathway associated with the first responder \u00df cells . Overall design: Ex vivo photoconversion of ins:CaMPARI islets and Fluorescence activated cell sorting FACS The primary islets from 4 mpf Tgins:CaMPARI animals were dissected from the fish post euthanasia.  Each islet was placed in a well of 24 well plate with 5 ml of DPBS medium supplemented with calcium  magnesium ThermoFisher  14040141  along with the appropriate concentration of D Glucose.  The islets were allowed to equilibrate for 5 min.  Subsequently  the entire 24 well plate was photoconverted using a 405 nm LED array Loctite  1167593 for 3 seconds.  The LED array evenly illuminated the entire dish with an intensity of 400 mW/cm2 and was triggered using a foot pedal switch.  An internal timer using an LED controller Loctite  1359255 was utilized for precise timing of light exposure.  The photoconverted islets were dissociated into single cells by incubation in TrypLE ThermoFisher  12563029 at 37\u00b0C in a benchtop shaker set at 450 rpm for 45 min.  Following dissociation  TrypLE was inactivated with 10% FBS  and the cells pelleted by centrifugation at 500 g for 10 min at 4\u00b0C.  The supernatant was carefully discarded  and the pellet re suspended in 500 \u00b5l of HBSS without xxx+  Mg2+.  To remove debris  the solution was passed over a 30 \u00b5m cell filter Miltenyi Biotec  130 041 407.  To remove dead cells  calcein violet ThermoFisher  C34858 was added at a final concentration of 1 \u00b5M and the cell suspension incubated at room temperature for 20 minutes.  The single cell preparation was sorted through 100 \u00b5m nozzle.  The following channels were utilized in FACS: Pacific Blue for calcein violet; FITC Fluorescein isothiocyanate for green fluorescence; and PE Phycoerythrin for red fluorescence.  Single cell RNA Sequencing For single cell RNA Seq  \u00df cells isolated from Tgins:CaMPARI islets were subjected to profiling using Smart Seq2 protocol from Takara.  For this  islets were isolated  photoconverted ex vivo  dissociated and FAC sorted as described above.  \u00df cells were collected from two conditions: responsive \u00df cells displaying photoconversion at 2.5 mM; and non responsive \u00df cells without xxx at 16.5 mM.  Photoconversion was defined as a high red to green ratio.  For FACS of each condition  index sorting was utilized  and the cells were collected in 96 well plate containing lysis buffer 2 \u00b5l of nuclease free water with 0.2% Triton X 100 and 4 U murine RNase Inhibitor NEB.  For each condition a total of 93 cells were collected for profiling.  Preparation of cDNA library and sequencing was performed exactly as described in a previous publication Singh et al.  Scientific Reports  2018; doi: 10.1038/s41598 018 35218 5.", null, "pubmed:38924394", null, "ExVi16Gr 02 E10", "GSM7655584", null, "source name:pancreas|tissue:pancreas|age:4 mpf|cell type:beta cell|genotype:Tgins:CaMPARI|treatment:photoconversion|geo loc name:missing|collection date:missing", "ExVi16Gr 02 E10", "Raw reads in fastq format were trimmed using trim galore with default parameters to remove adapter sequences.  Trimmed reads were aligned to the zebrafish genome  GRCz10  using HISAT2 with default parameters. htseq count was used to assign reads to exons thus eventually getting counts per gene. For analysis  cells with more than 800 detected genes were utilized.  Transcriptional profile of responsive and non responsive were compared using ROTS with default parameters.  Genes with false discovery rate FDR less than 0.05 were considered as differentially expressed. For gene ontology GO analysis of differentially expressed genes  FishEnrichR was utilized.  As background  genes expressed in at least 5 % of the cells were used. Assembly: GRCz10 Supplementary files format and content: csv", "pancreas", "The primary islets from 4 mpf Tgins:CaMPARI animals were dissected from the fish post euthanasia.  Each islet was placed in a well of 24 well plate with 5 ml of DPBS medium supplemented with calcium  magnesium ThermoFisher  14040141  along with the appropriate concentration of D Glucose.  The islets were allowed to equilibrate for 5 min.  Subsequently  the entire 24 well plate was photoconverted using a 405 nm LED array Loctite  1167593 for 3 seconds.  The LED array evenly illuminated the entire dish with an intensity of 400 mW/cm2 and was triggered using a foot pedal switch.  An internal timer using an LED controller Loctite  1359255 was utilized for precise timing of light exposure.", "The photoconverted islets were dissociated into single cells by incubation in TrypLE ThermoFisher  12563029 at 37\u00b0C in a benchtop shaker set at 450 rpm for 45 min.  Following dissociation  TrypLE was inactivated with 10% FBS  and the cells pelleted by centrifugation at 500 g for 10 min at 4\u00b0C.  The supernatant was carefully discarded  and the pellet re suspended in 500 \u00b5l of HBSS without xxx+  Mg2+.  To remove debris  the solution was passed over a 30 \u00b5m cell filter Miltenyi Biotec  130 041 407.  To remove dead cells  calcein violet ThermoFisher  C34858 was added at a final concentration of 1 \u00b5M and the cell suspension incubated at room temperature for 20 minutes.  The single cell preparation was sorted through 100 \u00b5m nozzle.  The following channels were utilized in FACS: Pacific Blue for calcein violet; FITC Fluorescein isothiocyanate for green fluorescence; and PE Phycoerythrin for red fluorescence. For single cell RNA Seq  \u03b2 cells isolated from Tgins:CaMPARI islets were subjected to profiling using Smart Seq2 protocol from Takara.  For this  islets were isolated  photoconverted ex vivo  dissociated and FAC sorted as described above.  \u03b2 cells were collected from two conditions: responsive \u03b2 cells displaying photoconversion at 2.5 mM; and non responsive \u03b2 cells without xxx at 16.5 mM.  Photoconversion was defined as a high red to green ratio.  For FACS of each condition  index sorting was utilized  and the cells were collected in 96 well plate containing lysis buffer 2 \u00b5l of nuclease free water with 0.2% Triton X 100 and 4 U murine RNase Inhibitor NEB.  For each condition a total of 93 cells were collected for profiling.  Preparation of cDNA library and sequencing was performed exactly as described in a previous publication Singh et al.  Scientific Reports  2018; doi: 10.1038/s41598 018 35218 5. Smart Seq2 from Takara", "4 mpf Tgins:CaMPARI zebrafish", "tissue:pancreas|age:4 mpf|cell type:beta cell|genotype:Tgins:CaMPARI|treatment:photoconversion", "GSM7655584", "GSM7655584: ExVi16Gr 02 E10; Danio rerio; RNA Seq", "GSM7655584 r1", "GSM7655584", "1", "The photoconverted islets were dissociated into single cells by incubation in TrypLE ThermoFisher  12563029 at 37\u00b0C in a benchtop shaker set at 450 rpm for 45 min.  Following dissociation  TrypLE was inactivated with 10% FBS  and the cells pelleted by centrifugation at 500 g for 10 min at 4\u00b0C.  The supernatant was carefully discarded  and the pellet re suspended in 500 \u00b5l of HBSS without xxx+  Mg2+.  To remove debris  the solution was passed over a 30 \u00b5m cell filter Miltenyi Biotec  130 041 407.  To remove dead cells  calcein violet ThermoFisher  C34858 was added at a final concentration of 1 \u00b5M and the cell suspension incubated at room temperature for 20 minutes.  The single cell preparation was sorted through 100 \u00b5m nozzle.  The following channels were utilized in FACS: Pacific Blue for calcein violet; FITC Fluorescein isothiocyanate for green fluorescence; and PE Phycoerythrin for red fluorescence. For single cell RNA Seq  \u03b2 cells isolated from Tgins:CaMPARI islets were subjected to profiling using Smart Seq2 protocol from Takara.  For this  islets were isolated  photoconverted ex vivo  dissociated and FAC sorted as described above.  \u03b2 cells were collected from two conditions: responsive \u03b2 cells displaying photoconversion at 2.5 mM; and non responsive \u03b2 cells without xxx at 16.5 mM.  Photoconversion was defined as a high red to green ratio.  For FACS of each condition  index sorting was utilized  and the cells were collected in 96 well plate containing lysis buffer 2 \u00b5l of nuclease free water with 0.2% Triton X 100 and 4 U murine RNase Inhibitor NEB.  For each condition a total of 93 cells were collected for profiling.  Preparation of cDNA library and sequencing was performed exactly as described in a previous publication Singh et al.  Scientific Reports  2018; doi: 10.1038/s41598 018 35218 5. 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