|
| Status |
Public on Jan 03, 2017 |
| Title |
co-culture treated with Bic/4AP for 1 h Hs+Ms_Bic-4AP_1h_2 |
| Sample type |
SRA |
| |
|
| Source name |
hiPSC-derived neurons
|
| Organisms |
Homo sapiens; Mus musculus |
| Characteristics |
cell type: forebrain neuron-like, heterogenous
passages: from iPSC passages 30-40
iPSc line: iPS D1
developmental stage: postmitotic
|
| Treatment protocol |
Bicuculline (50 µM) applied together with 4-aminopyridine (250 µM) (Bic/4AP)
|
| Growth protocol |
We differentiated iPSCs to NPCs using a slightly modified spin embryoid body protocol (Kim et al., 2011) with dual SMAD inhibition (Chambers et al., 2009). Differentiation of NPCs was initiated by plating 50,000 cells/cm2 on PO/Lam dishes without EGF and FGF2. Neuronal differentiation was allowed to proceed for 7 (co-cultures) or 10 (hiPSCd neuron-only cultures) weeks.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was purified from cultured cells (3.5 cm dish) with RNeasy Mini (hiPSCd neuron/mouse primary neuron co-cultures) or Micro (hiPSCd neuron-only cultures) Kit (Qiagen) along with on-column RNase-Free DNase (Qiagen) digestion of DNA
Poly(A)+ RNA was isolated from total RNA with the NEBNext Poly(A) mRNA Magnetic Isolation Module (New England Biolabs). cDNA libraries were prepared with NEBNext Ultra Directional RNA Library Prep Kit for Illumina (New England Biolabs). NEBNext Multiplex Oligos for Illumina (New England Biolabs) were used.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
P0 mouse primary neurons were added to hiPSCd neurons for the last 10 days of differentiation.
polyA purified
|
| Data processing |
Illumina Casava1.7 software used for basecalling.
hiPSCd neuron/mouse primary neuron co-culture sample reads were filtered for human reads as follows. Paired-end reads were aligned independently, without pairing, to the mouse genome assembly GRCm38/mm10 with Bowtie2 version 2.1.0. Unaligned read pairs were joined and split again to eliminate pairs with one aligned read. Then, genome-unaligned paired-end reads were aligned to GRCm38/mm10 transcriptomes (UCSC Genes/knownGene), respectively, with Bowtie2 (2.1.0). The reads not aligned to the mouse genome and transcriptome were handled as reads obtained from hiPSCd neuron-only samples.
Processed mixed-species sample reads and human-cells-only sample reads were aligned to gene model annotations of the human genome assembly GRCh37/hg19 with TopHat version v2.0.9.
Mapped reads per gene were counted strand-specifically with HTSeq-count (HTSeq version 0.6.0 or 0.6.1) in the union overlap resolution mode.
Differentially expressed genes were determined with the DESeq2 package version 1.8.1.
Genome_build: GRCh37/hg19
Supplementary_files_format_and_content: HTSeq-count read counts per gene.
|
| |
|
| Submission date |
Oct 14, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Priit Pruunsild |
| E-mail(s) |
pruunsild@nbio.uni-heidelberg.de
|
| Organization name |
University of Heidelberg
|
| Department |
Neurobiology
|
| Street address |
INF 364
|
| City |
Heidelberg |
| ZIP/Postal code |
69120 |
| Country |
Germany |
| |
|
| Platform ID |
GPL16512 |
| Series (1) |
| GSE88773 |
Networks of cultured iPSC-derived neurons reveal the human synaptic activity-regulated adaptive gene program |
|
| Relations |
| BioSample |
SAMN05908866 |
| SRA |
SRX2245892 |
