characteristics.text: 2013-02-08T17:03:00Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T16:09:43Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA1712779
characteristics.characteristic: External Id
characteristics.text: Pediatric Diabetes Research Center, UCSD Stem Cell Program
characteristics.characteristic: INSDC center name
characteristics.text: 2013-02-08T17:03:00Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T16:09:43Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS197535
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-1086:HI41
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: islet cell
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/UBERON_0000016
characteristics.characteristic: cell type
characteristics.text: human
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: common name
characteristics.text: Protocols: Briefly, cells were lysed in Trizol reagent (Invitrogen) for extraction of total RNA. Residual contaminating genomic DNA was removed using the Turbo Dnase kit (Ambion). mRNA was isolated from 2 ug of DNA-free total RNA using the Dynabeads mRNA Purification kit (Invitrogen). Strand-specific RNA-seq libraries were prepared as previously described [Parkhomchuk, D., et al., Nucleic Acids Res, 2009. 37:e123], with minor modifications. Following purification, mRNA was primed with Olig(dT)s and random hexamers, and reverse-transcribed to first-strand cDNA. Residual dNTPs were removed from the first-strand cDNA using Illustra MicroSpin G-25 columns (GE Healthcare). In the second-strand synthesis reaction, dUTPs were used instead of dTTPs. The converted double-strand cDNA was fragmented using a Bioruptor Sonicator for 60 cycles of 30 sec on and 30 sec off. After end-repair and adenine base addition, the cleaved double-strand cDNA fragments were ligated to Pair-end Adaptor Oligo Mix (Illumina) and size-fractionated on a 2% high resolution agarose gel (Sigma). cDNA fragments of 200 (+25 bp,-25bp) were recovered, and incubated with uracil-N-glycosylase (UNG) to digest the second-strand cDNA. Purified single-strand cDNA was then used as template for 15 cycles of amplification using pair-end PCR primers (Illumina). The amplified products were separated on a 2% agarose gel and a band between 225-275 bp excised.
characteristics.characteristic: description
characteristics.text: adult
characteristics.characteristic: developmental stage
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_9606
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: HI41
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0006980
characteristics.characteristic: title