|
Status |
Public on Mar 26, 2014 |
Title |
gfp_nt_1hr_3 |
Sample type |
SRA |
|
|
Source name |
GFP positive cells from 5 dpf larvae
|
Organism |
Danio rerio |
Characteristics |
drug: no drug time: 1hr gfp status: positive genotype: Tg(sqET20) experiment_date: Mar2012
|
Treatment protocol |
For neomycin treatment, 5dpf larvae were treated for 30min with 300μM neomycin (Fisher BioReagents) diluted in 0.5X E2 medium, rinsed three times and recovered in 0.5X E2 medium at 28.5°C for the indicated time.
|
Growth protocol |
Larvae were generated by paired matings and raised in 0.5X E2 medium at 28.5°C. GFP positive larvae were sorted from wild-type larvae at 48 hpf under fluorescent stereoscope.
|
Extracted molecule |
total RNA |
Extraction protocol |
Two hundred 5dpf untreated Tg(sqET20) control or neomycin treated Tg(sqET20) larvae were anesthetized, collected in a 2mL tube, dissociated with trypsin and filtered to remove un-dissociated tissue. A two-gate FACS strategy was used to select only living target cells from excesses of dead cells and cellular debris of the larval cell suspensions (see Materials and Methods of associated manuscript for detailed FACS protocol). Approximately 30,000 GFP- or GFP+ cells were used for total RNA extraction using Trizol (Invitrogen) following the manufacturer’s manual. Libraries for RNA sequencing were made with poly-A selected mRNA using the Illumina TruSeq RNA library construction kit v2 (Illumina). The resulting libraries were purified using Agencourt AMPure XP system (Backman Coulter), then quantified using a Bioanalyzer or Qubit Fluorometer (Life Technologies).
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|
|
Library strategy |
RNA-Seq |
Library source |
transcriptomic |
Library selection |
cDNA |
Instrument model |
Illumina HiSeq 2000 |
|
|
Data processing |
Data was processed with CASAVA version 1.8.2 Sequence reads in fastq format were mapped to danRer7using tophat-1.4.1 with options –g 1 and a GTF description of transcripts based on Ensembl 63. FPKM values for these transcripts were generated using cufflinks v1.3. Genome_build: danRer7 Supplementary_files_format_and_content: FPKM values per gene.
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|
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Submission date |
Mar 25, 2014 |
Last update date |
May 15, 2019 |
Contact name |
Chris W Seidel |
E-mail(s) |
seidel@phageT4.org
|
Phone |
816 926 9054
|
Organization name |
Stowers Institute
|
Department |
Genomics
|
Lab |
Seidel
|
Street address |
1000 E 50th St
|
City |
Kansas City |
State/province |
MO |
ZIP/Postal code |
64110 |
Country |
USA |
|
|
Platform ID |
GPL14875 |
Series (1) |
GSE56176 |
Gene expression analysis of hair cell regeneration in the zebrafish lateral line |
|
Relations |
BioSample |
SAMN02700067 |
SRA |
SRX501289 |