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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Nov 01, 2011 |
| Title |
mouse feces NM1200_9.36 |
| Sample type |
SRA |
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| Source name |
mouse feces
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| Organisms |
Bacteroides; Ruminococcus; Streptococcus thermophilus; Lactobacillus; Collinsella; Clostridia; Parabacteroides; Bifidobacterium animalis subsp. lactis CNCM I-2494 |
| Characteristics |
sample type: fecal pellet of a mouse
source: the fecal pellet of a mouse (C57Bl/6J) colonized with 15 species of gut bacteria (B. caccae, B. ovatus, B. thetaiotaomicron, B. uniformis, B. vulgatus, B. WH2, C. aerofaciens, C. scindens, C. spiroforme, D. longicatena, E. rectale, F. prausnitzii, P. distasonis, R. obeum, R. torques) 36 days prior which was also gavaged with 5 fermented milk strains (B. animalis subsp. lactis, L. lactis subsp. cremoris, two strains of L. delbrueckii subsp. bulgaricus, S. thermophilus) 1, 14, 15, 21, and 22 days prior. Mouse was fed B&K rat and mouse autoclavable chow #7378000, Zeigler Bros., Inc.).
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| Extracted molecule |
total RNA |
| Extraction protocol |
Libraries were prepared according a slightly modified version of the protocol accompanying the Illumina Genomic DNA Sample Prep Kit. Briefly, cDNA was sonicated in a BioRuptor XL water bath sonicator, cleaned up and concentrated using a Qiagen PCR Purification column, and end-repaired using Klenow DNA polymerase. The blunt DNA was treated with Klenow fragment (exo minus) to add an adenine overhang, and the A-tailed molecules were ligated to the relevant Illumina adapter sequence (with or without a 4-nt barcode). Adaptered-DNA was then size-selected by agarose gel electrophoresis. Fragments of the appropriate size were PCR amplified and purified, after which the purified PCR products were loaded on an Illumina flow cell for cluster generation. Libraries were sequenced on the Genome Analyzer (IIx) following the manufacturer's protocols.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer IIx |
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| Description |
Bacteroides caccae ATCC 43185,Bacteroides ovatus ATCC 8483,Bacteroides thetaiotaomicron VPI-5482,Bacteroides uniformis ATCC 8492,Bacteroides vulgatus ATCC 8482,Bacteroides WH2,Bifidobacterium animalis subsp. lactis CNCM I-2494,Clostridium scindens ATCC 35704,Clostridium spiroforme DSM 1552,Collinsella aerofaciens ATCC 25986,Dorea longicatena DSM 13814,Eubacterium rectale ATCC 33656,Faecalibacterium prausnitzii M21/2,Lactobacillus delbrueckii subsp. bulgaricus CNCM I-1632,Lactobacillus delbrueckii subsp. bulgaricus CNCM I-1519,Lactococcus lactis subsp. cremoris CNCM I-1631,Parabacteroides distasonis ATCC 8503,Ruminococcus obeum ATCC 29174,Ruminococcus torques ATCC 27756,Streptococcus thermophilus CNCM I-1630
rRNA-depleted total RNA
RNA isolated from the fecal pellet of a mouse (C57Bl/6J) colonized with 15 species of gut bacteria (B. caccae, B. ovatus, B. thetaiotaomicron, B. uniformis, B. vulgatus, B. WH2, C. aerofaciens, C. scindens, C. spiroforme, D. longicatena, E. rectale, F. prausnitzii, P. distasonis, R. obeum, R. torques) 36 days prior which was also gavaged with 5 fermented milk strains (B. animalis subsp. lactis, L. lactis subsp. cremoris, two strains of L. delbrueckii subsp. bulgaricus, S. thermophilus) 1, 14, 15, 21, and 22 days prior. Mouse was fed B&K rat and mouse autoclavable chow #7378000, Zeigler Bros., Inc.).
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| Data processing |
After parsing each sequencing lane's data by barcode (where appropriate), we excised the barcodes and mapped the reads to an appropriate set of reference genomes using the ssaha2 algorithm. Minimum score thresholds for ssaha were selected based on the distribution of scores for all mapped reads of a 32-nt barcoded sample and a 36-nt non-barcoded sample (29 was selected as the minimum score for 32-nt barcoded samples; 33 was the minimum score used for 36-nt and 76-nt non-barcoded samples). Although an 18-nt read is sufficient to map more than 90% of the sequencing reads, even at 32-36 nucleotides a large fraction of reads map to multiple locations (within a genome and/or across genomes). Reads that map non-uniquely were added to each gene in proportion to each gene's fraction of unique-match counts (e.g., a non-unique read that maps equally well to gene A with 18 unique reads and gene B with 2 unique reads will result in 0.9 counts being tallied to gene A and 0.1 counts being tallied to gene B). We added a pseudocount (i.e. added a value of 1) to each gene count prior to normalization to account for differences in sampling depth, although clearly a more appropriate model-based approach to smooth lower expression values is needed in the future, particularly with mixed species samples where one or more species is clearly undersampled. Raw counts were ultimately normalized to reads/kb gene length/million mapped reads (RPKM) following gene count assignments.
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| Submission date |
Aug 25, 2011 |
| Last update date |
May 15, 2019 |
| Contact name |
Nathan P McNulty |
| E-mail(s) |
nathan.p.mcnulty@gmail.com
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| Phone |
314-362-3963
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| Organization name |
Washington University School of Medicine
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| Department |
Center for Genome Sciences and Systems Biology
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| Lab |
Gordon
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| Street address |
4444 Forest Park Ave. (5th Floor)
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| City |
Saint Louis |
| State/province |
MO |
| ZIP/Postal code |
63108 |
| Country |
USA |
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| Platform ID |
GPL14349 |
| Series (1) |
| GSE31670 |
The impact of a consortium of fermented milk strains on the gut microbiome of gnotobiotic mice and monozygotic twins. (RNA-Seq) |
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| Relations |
| SRA |
SRX093143 |
| BioSample |
SAMN00714075 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM786165_NM1200_9.36.counts.txt.gz |
205.3 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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