 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jan 21, 2024 |
| Title |
Biol rep 1 CSPG-negative cells |
| Sample type |
SRA |
| |
|
| Source name |
Hindbrain St, 18
|
| Organism |
Gallus gallus |
| Characteristics |
tissue: Hindbrain St, 18
cell type: Neural ectoderm
isolation: FACS isolation for CSPG-
|
| Growth protocol |
For each biological replicate, 30 hindbrains of st. 18 HH chick embryos were dissected and separated into single cells using collagenase type IV (200 Units\ml, Worthington, 47B9407). Cells were washed in PBS, centrifuged and incubated with mouse anti-CSPG antibody for 75 min (1:50, (#c8053; Sigma-Aldrich), diluted in MACSn MACS® BSA Stock Solution and autoMACS® Rinsing Solution (1:20, Miltenyi Biotec), washed in PBS, centrifugaed, and incubated with anti-mouse Alexa 488 antibody (1:200, Thermo-Fishe) diluted in autoMACS® Running Buffer (Miltenyi Biotec). Cells were washed in PBS , centrifuged and sortes using ARIA III FACS for CSPG-positive and CSPG-negative cell groups.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
mRNA was extracted from the FACS -isolated CSPG-positive and CSPG-negative cells using Single Cell RNA purification kit (Norgen Biotek Corp). Each RNA sample had a RIN >6.9.
Libraries were prepared using the KAPA Stranded mRNA-Seq Kit (KR0960, Roche, USA) and Illumina platforms sample preparation protocol (v3.15) according to manufacturer's protocoal. RNA sequencing was conducted by Illumina NextSeq 2000 machine using NextSeq 2000 P2, 100 cycles kit (Illumina, USA). The output was ~25 million single-end- 120bp reads per sample.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
NextSeq 2000 |
| |
|
| Description |
processed data columns: A-I, general gene details; J-S, raw counts per sample; T-AC, normalized counts per sample; AD, average signal over all samples; AE-AI, DESeq2 results for comparing CSPG-positive cells to CSPG-negative cells
|
| Data processing |
Fastq files were created with bcl2fastq v2.20.0.422. Cutadapt, v3.4, was used for removal of adapters, polyA and low-quality sequences.
Alignment to GRCg6a (chicken) was done with TopHat, v2.1.1, with annotations from Ensembl release 99. Quantification was done with htseq-count, v0.13.5.
Differential gene expression analysis was performed using the R package DESeq2, v1.30.0. Significance threshold was taken as padj<0.05.
Assembly: GRCg6a
Supplementary files format and content: Excel file with general gene details, raw and normalized counts, and the results of comparing CSPG-positive cells to CSPG-negative cells with DESeq2.
|
| |
|
| Submission date |
Apr 27, 2023 |
| Last update date |
Jan 21, 2024 |
| Contact name |
Yuval Nevo |
| Organization name |
The Hebrew University of Jerusalem
|
| Lab |
Info-CORE
|
| Street address |
Ein Kerem Campus
|
| City |
Jerusalem |
| ZIP/Postal code |
9112102 |
| Country |
Israel |
| |
|
| Platform ID |
GPL32831 |
| Series (1) |
| GSE230804 |
The neural-progenitor/stem-cell (NPSC) properties of hindbrain boundary cells |
|
| Relations |
| BioSample |
SAMN34411215 |
| SRA |
SRX20121219 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
|
|
|
|
 |
