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| Status |
Public on Dec 16, 2021 |
| Title |
C5 |
| Sample type |
SRA |
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| Source name |
control_isolated retinal microglia
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| Organism |
Rattus norvegicus |
| Characteristics |
strain: Dark Agouti
treatment: citrate buffer control
blood glucose: normal
cell type: isolated retinal microglia
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| Treatment protocol |
Rats received a dose of steptozotocin (STZ, 55mg/kg) or citrate buffer control at 10am after 12 hours of fasting. Blood glcuose levels were measured 24 hours later to check for hyperglycaemia. After 1 week blood glucose was measured bi-weekly and diabetic rats were treated with 2 Units of Protaphane (insulin) if blood glucose reaadings were >30mmol/l. Animals were taken out to 4 weeks post STZ-treatment.
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| Growth protocol |
Rats were rendered diabetic at 8-10 weeks of age
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| Extracted molecule |
total RNA |
| Extraction protocol |
Whole retina from control and 4 week STZ-treated rats (n = 5 control, n = 4 STZ) were isolated, papain digested (Worthington Biochemical, NJ, USA), and labelled with CD11b-FITC conjugate (Miltenyi Biotec, Bergisch Gladbach, Germany) and microglia were isolated via FACS. RNA was extracted from isolated populations (RNeasy Micro Kit, Qiagen, Hilden, Germany) and purity analysed (Agilent Technologies, Santa Clara, CA, USA). A SmartSeq v4 kit (Clontech Laboratories, Mountain View, CA, USA) was used for a preamplification step.
RNA libraries were prepared for sequencing using standard Illumina protocols - NexteraXT
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
C5_CAF4FANXX_GGACTCCT_L007
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| Data processing |
Image analysis was performed in real time by the HiSeq Control Software (HCS) v2.2.68 and Real Time Analysis (RTA) v1.18.66.3, running on the instrument computer. RTA performs real-time base calling on the HiSeq instrument computer. Then the Illumina bcl2fastq 2.18.0.12 pipeline was used to generate the sequence data.
The reads were also screened for the presence of any Illumina adaptor/overrepresented sequences and cross-species contamination. The cleaned sequence reads were then aligned against the genome of Rattus norvegicus (Ensemble verison Rnor_6.0 (GCA_000001895.4)).
Tophat aligner (v2.0.14) was used to map reads to the genomic sequences and are in the compressed BAM format
The transcripts were assembled with the Stringtie tool v1.2.4 utilising the reads alignment and reference annotation based assembly option (RABT).
Differential expression analysis was carried out using the bioconductor package edgeR. Default normalisation method (TMM) was used.
Genome_build: Rnor_6.0
Supplementary_files_format_and_content: gene counts, transcript assemblies with TPM/FPKM values, differential expression analysis table
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| Submission date |
Oct 23, 2019 |
| Last update date |
Dec 16, 2021 |
| Contact name |
Andrew Ian Jobling |
| E-mail(s) |
aij@unimelb.edu.au
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| Organization name |
University of Melbourne
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| Department |
Anatomy & Neuroscience
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| Street address |
Medical Building, Cnr Grattan St and Royal Pde
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| City |
Melbourne |
| State/province |
Victoria |
| ZIP/Postal code |
3010 |
| Country |
Australia |
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| Platform ID |
GPL18694 |
| Series (1) |
| GSE139276 |
RNAseq analysis of retinal microglia during early diabetes |
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| Relations |
| BioSample |
SAMN13091491 |
| SRA |
SRX7044068 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4135927_C5_CAF4FANXX_GGACTCCT_L007_R1.fastq.transcripts.gtf.gz |
3.6 Mb |
(ftp)(http) |
GTF |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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