characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2017-08-29T17:02:00Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2017-03-30T13:18:52Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA103950299
characteristics.characteristic: External Id
characteristics.text: National Research Laboratory of Molecular Microbiology and Toxicology, Department of Agricultural Biotechnology, Center for Food Safety and Toxicology, Seoul National University
characteristics.characteristic: INSDC center name
characteristics.text: 2017-08-29T17:02:00Z
characteristics.characteristic: INSDC first public
characteristics.text: 2017-03-30T13:18:52Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS1639467
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-5629:HT29-MTX EXPOSURE1
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: Protocols: V. vulnificus MO6-24/O cells (approximately 2 X 10^8 colony forming unit) exposed to HT29-methotrexate cells (2 X 10^7 cells in the cell culture dish (Nunc)) for 2 h were fixed with 2:1 volumes of RNAprotect Bacteria Reagent (Qiagen) and harvested. Total RNAs were isolated from the V. vulnificus cells using miRNeasy Mini Kit (Qiagen). Genomic DNA in the purified RNA was removed with digestion using TURBO^TM DNase (Ambion) to below PCR-detectable levels. RNA quality was determined using Bioanalyser (Agilent) 23S and 16S rRNA of total RNAs were depleted using MICROBExpress kit (Ambion). The enriched mRNAs were fragmented using Fragment, Prime, Finish Mix (Illumina). The mixtures were reacted at 94°C for 8 min and then incubated at 4°C. Synthesis of first strand cDNA was prepared by mixing 0.5 μg of the enriched mRNA, First Strand Synthesis Act D Mix (Illumina), and Superscript® II polymerase (Invitrogen). The mixture was incubated at 25°C for 10 min and at 42°C for 45 min and the polymerase was finally inactivated at 75°C for 15 min. Remaining dNTPs were removed AMPure XP beads. Then, the second strand of cDNA was further synthesized using Second Strand Marking Master (Illumina). The mixture was incubated at 16°C for 60 min. Remaining dUNTPs were removed AMPure XP beads. A single 3’ adenosine moiety was added to the cDNA fragments using A-Tailing mix (Illumina) The Illumina adapters (containing primer sites for sequencing and flowcell surface annealing) were ligated onto the repaired ends on the cDNA fragments using Ligation Mix (Illumina). The mixtures were incubated at °C for 10 min and then add the Stop Ligation Buffer (Illumina) for inactivation of ligation. Final mixtures were purified using AMPure XP beads.
characteristics.characteristic: description
characteristics.text: exposed to HT29-methotrexate cells for 2 h
characteristics.characteristic: growth condition
characteristics.text: Vibrio vulnificus
characteristics.characteristic: organism
characteristics.text: Vibrio vulnificus
characteristics.characteristic: organism
characteristics.text: Vibrio vulnificus
characteristics.characteristic: scientific_name
characteristics.text: MO6-24/O
characteristics.characteristic: strain
characteristics.text: HT29-MTX EXPOSURE1
characteristics.characteristic: title