characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2023-04-01T00:23:05Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2023-04-01T00:23:05Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA110461477
characteristics.characteristic: External Id
characteristics.text: Latvian Biomedical Research and Study Centre
characteristics.characteristic: INSDC center name
characteristics.text: 2023-04-01T00:23:05Z
characteristics.characteristic: INSDC first public
characteristics.text: 2023-04-01T00:23:05Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS12559505
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-12013:GEV-013_F1
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.characteristic: broker name
characteristics.text: human
characteristics.characteristic: common name
characteristics.text: Protocols: Feces samples were stored at -80 C. For EVs isolation samples were treated and concentrated. In brief, 2 grams of the thawed aliquot was suspended in 8 ml PBS-DEPC with vigorous vortex until the sample is homogeneous. Samples were centrifuged once 1000 × rcf for 10 min, once- 2000 × rcf for 10 min and 3-5 times 3200 × rcf for 10 min at +4C until the supernatant is transparent. The sediment was get rid of and the tubes were changed after each centrifugation step. The final supernatant was diluted 1:1 with PBS-DEPC and filter, first, through 0,45 ul filter , than through 0,20 ul filter (Millipore). Filtered samples were tranfered into Amicon® Ultra-15 Centrifugal Filter Units and centrifuged at +4C x3200 rcf for 10 mins as many times as to get 1 ml of concentrated sample. Thus, samples were ready for loading on EVs isolation column. Before RNA extraction, the EV samples were treated with 1 mg/ml proteinase K (Thermo Fisher Scientific, USA) for 30 min at + 37˚C. Proteinase was inactivated by heating the sample at + 65˚C for 10 minutes, and then the samples were treated with 10 ng/μl RNAse A (Thermo Fisher Scientific, USA) for 15 minutes at +37˚C. Immediately after that, EV samples were lysed by adding 5 volumes of QIAzol Lysis reagent and EV-RNA was extracted using miRNeasy Micro Kit (Qiagen, USA) according to the manufacturer’s protocol. Briefly, 1 ml of chloroform was added to the EV lysate, incubated at room temperature for 10 minutes and centrifuged for 15 minutes at 12000 g at +4˚C. The aqueous phase was transferred to fresh tube and 1.5 vol of 100% ethanol was added, and the mixture was loaded onto MinElute Spin column. Columns were centrifuged and washed according to manufacturer’s suggestions and the RNA was eluted using 12 μl of RNAse-free water. To determine the concentration and assess the quality, the RNA was measured using Agilent 2100 Bioanalyzer and RNA 6000 Pico Kit (Agilent technologies, USA).From the tissue specimens, approximately 20 mg of tissues were homogenized using QIAzol Lysis Reagent (Qiagen, USA) and Lysing Matrix A tubes in a FastPrep-24 homogenizer (MP Biomedicals, USA). RNA was isolated using miRNeasy Micro Kit (Qiagen) following the small RNA enrichment protocol provided by the manufacturer. EV-RNA libraries were constructed from half of the entire yield of extracted EV RNA without any size separation steps, whereas tissue small RNA libraries were constructed from 10 ng of tissue small RNA fraction using CleanTag® Small RNA Library Prep Kit (Trilink Biotechnologies, USA). The libraries were cleaned using Blue Pippin DNA Size Selection with 3% gel Blue Pippin Cassette (Sage Science, USA) setting target length to 130 - 250 bp. Transcriptome libraries were built from 100 ng of long tissue RNA fraction using TruSeq Stranded mRNA library Prep (Illumina, USA) following manufacturer's instructions. Libraries were size-selected using Blue Pippin system with 2% gel Blue Pippin Cassette (Sage Science, USA) with a range of 200 - 600 bp and their quality and quantity was assessed using Agilent DNA kit (Agilent technologies, USA).
characteristics.characteristic: description
characteristics.text: adult
characteristics.characteristic: developmental stage
characteristics.text: gastric cancer
characteristics.characteristic: disease
characteristics.text: extracellular vesicles
characteristics.characteristic: fraction
characteristics.text: GEV-013
characteristics.characteristic: individual
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: Homo sapiens
characteristics.characteristic: organism
characteristics.text: feces
characteristics.characteristic: organism part
characteristics.text: Homo sapiens
characteristics.characteristic: scientific_name
characteristics.text: GEV-013_F1
characteristics.characteristic: title