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Sample GSM5526514 Query DataSets for GSM5526514
Status Public on Jun 29, 2022
Title 119
Sample type SRA
 
Source name pancreatic islet
Organism Mus musculus
Characteristics cell type: Pancreatic islets
Sex: male
strain background: C57BL6/J
diet: Western diet
genotype: Zfp148floxed/floxed
age (wks): 22
Treatment protocol Mice were kept on a high-fat/high-sucrose Western-style diet (Envigo Teklad TD.08811, IACUC protocol A005821-R01) for 20 weeks.
Growth protocol β-Zfp148KO and control littermate mice of both sexes were used.
Extracted molecule total RNA
Extraction protocol Mouse islets were isolated as described (Rabaglia et al. American Journal of Physiology-Endocrinology and Metabolism 289, E218-E224 (2005). PMID: 15741243). After isolation, islets were placed in 350 uL of QIAzol Lysis Reagent (Qiagen) and stored at -80˚C before purifying RNA using Mini RNeasy purification kit. Purified RNA was reverse-transcribed and sequenced by the UW Madison Biotechnology Center.
RNA libraries were prepared for sequencing at the UW Madison Biotechnology Center. RNA integrity was determined using assayed on Agilent 2100 BioAnalyzer. Libraries were prepared using Illumina® TruSeq® Stranded Total RNA Sample Prep Gold kit (Illumina Inc., San Diego, California, USA). Complementary DNA (cDNA) was purified using Agencourt AMPure XP bead purification (Beckman-Coulter) following rRNA reduction. Libraries were quantified using Qubit DNA HS kit (ThermoFisher) and assayed on Agilent HS DNA Chips. Sequencing was done on Illumina HiSeq 2500 units.
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2500
 
Description For generation of the mice, see Keller et al. JCI 2019.
Islet RNA sequencing
Data processing Mouse islet RNA-sequencing reads were mapped via bowtie against the RefSeq mm10 mouse reference genome.
Gene expression values were estimated via RSEM.
Expected transcripts per million were normalized using median-by-ratio normalization.
Normalized gene abundance was analyzed for differential expression by EBSeq (Leng, N., Dawson, JA., Stewart, RM., Ruotti, V., Rissman, AI., Smits, BMG., Haag, JD., Gould, MN., Thomson, JA., and Kendziorski, C. (2013). EBSeq: an empirical Bayes hierarchical model for inference in RNA-seq experiments. Bioinformatics 29.8 1035-1043). Significantly different genes were determined as having posterior probability of differential expression > 0.95 (equivalent to FDR < 0.05). For coding genes, PPDE was determine for three tests: Zfp148floxed/floxed vs beta-ZFP148KO males, Zfp148floxed/floxed vs beta-ZFP148KO females, Zfp148floxed/floxed vs beta-ZFP148KO with both sexes included. Only genes intersecting all of these tests with PPDE > 0.95 were included for further analysis. This was repeated for ncRNAs, and the resulting genes meeting PPDE threshold for coding and non-coding genes were pooled into the final list for GO and subsequent analyses.
Genome_build: mm10/GRCm38
Supplementary_files_format_and_content: Emfinger_et_al_RNA_seq_PPDE_0.95.xlsx: This is an Excel (.xlsx) file containing tabs for the PPDE results for the three independent tests for coding and non-coding sequences, and the final intersected list. Transcript abundance is indicated for each sample in transcript reads per million (TPM).
 
Submission date Aug 17, 2021
Last update date Jun 29, 2022
Contact name Alan Attie
E-mail(s) adattie@wisc.edu, attie@biochem.wisc.edu
Organization name University of Wisconsin-Madison
Department Biochemistry
Lab Attie lab
Street address 433 Babcock Drive
City Madison
State/province Wisconsin
ZIP/Postal code 53706
Country USA
 
Platform ID GPL17021
Series (1)
GSE182311 β-cell-specific deletion of Zfp148 improves nutrient-stimulated β-cell Ca2+ responses
Relations
BioSample SAMN20840906
SRA SRX11804477

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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