characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2018-04-16T17:02:47Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2017-03-02T11:20:50Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA103137418
characteristics.characteristic: External Id
characteristics.text: Department of Veterinary Population Medicine, University of Minnesota, Twin Cities.
characteristics.characteristic: INSDC center name
characteristics.text: 2018-04-16T17:02:47Z
characteristics.characteristic: INSDC first public
characteristics.text: 2017-03-02T11:20:50Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS1575631
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-5546:Placebo 2
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: Protocols: Stool samples were collected into 1.7ml RNase/DNase-free tubes (Catalog #: C-2170, Denville Scientific, Holliston, MA, USA) at time points accordingly. The fecal samples were immediately frozen on dry ice and then stored at -80 °C Mice received morphine and pellet implantation method as described(Bryant et al. 1988). Using this method, plasma levels of morphine are in the 0.6-2.0-microg/ml range (range seen in opioid abusers and patients on opioids for moderate to severe pain). Furthermore, this model is commonly used in the study of opiate dependence and addiction(Bryant et al. 1988). Briefly, placebo or 25 mg morphine were inserted in a small pocket created by a small skin incision on the animal’s dorsal side; incisions were closed using surgical wound clips (Stoelting, 9 mm Stainless Steel, Wooddale, IL). DNA extractions from fecal matters were carried out using the PowerSoil DNA isolation kit (Catalog #: 12888-100, MO BIO Laboratories, Carlsbad, CA, USA). All extracted DNA samples were stored at -80°C until amplification. Fecal DNA samples (25 ng) were used as template for PCR amplification of the V4 region of the 16S rRNA gene. Degenerate primer sets were designed with Illumina index sequences on the 5 end of the reverse primer, which were specific to each fecal DNA sample and allowed for multiplexed sequencing. Primers also contained Illumina PCR primer sequences (reverse primer) and Illumina TruSeq Universal Adapter sequences (forward primers) for library creation. 1) Primer Sequences (16S-specific portion in bold) Meta_V4_515F: TCGTCGGCAGCGTCAGATGTGTATAAGAGACAGGTGCCAGCMGCCGCGGTAA Meta_V4_806R: GTCTCGTGGGCTCGGAGATGTGTATAAGAGACAGGGACTACHVGGGTWTCTAAT The indexing primers are as follows. This step adds both the index and the flowcell adapters. [i5] and [i7] refer to the index sequence codes used by Illumina. The p5 and p7 flow cell adapters are in bold. Forward indexing primer: AATGATACGGCGACCACCGAGATCTACAC[i5]TCGTCGGCAGCGTC Reverse indexing primer: CAAGCAGAAGACGGCATACGAGAT[i7]GTCTCGTGGGCTCGG 2) Cycling Conditions -PCR reactions are performed using KAPA HiFidelity Hot Start Polymerase PCR 1 (using the Meta_V4_515F/Meta_V4_806R primer pair) 95C - 5 minutes 20 cycles of: 98C - 20 seconds 55C - 15 seconds 72C - 1 minute Hold at 4C -After the first round of amplification, PCR 1 products are diluted 1:100 and 5 ul of 1:100 PCR 1 is used in the second PCR reaction. PCR 2 (using different combinations of forward and reverse indexing primers) 95C - 5 minutes 10 cycles of: 98C - 20 seconds 55C - 15 seconds 72C - 1 minute Hold at 4C
characteristics.characteristic: description
characteristics.text: wild type genotype
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0005168
characteristics.characteristic: genotype
characteristics.text: Mus musculus domesticus
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_10092
characteristics.characteristic: host organism
characteristics.text: mouse gut metagenome
characteristics.characteristic: organism
characteristics.text: mouse gut metagenome
characteristics.characteristic: organism
characteristics.text: feces
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCIT_C13234
characteristics.characteristic: organism part
characteristics.text: mouse gut metagenome
characteristics.characteristic: scientific_name
characteristics.text: C57BL/6J
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0000606
characteristics.characteristic: strain
characteristics.text: Placebo 2
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0001674
characteristics.characteristic: title