|
| Status |
Public on Feb 10, 2017 |
| Title |
CACO-2 cells incubated with HMO-grown B. infantis, replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
CACO-2 cells_HMO-grown B. infantis
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: CACO-2
incubated with: HMO-grown B. infantis
|
| Growth protocol |
B. infantis ATCC 15697 from the exponentially grown 48h-old cultures supplemented with HMO, GLU or LAC were collected by centrifugation (4000g for 10 min), washed, and resuspended in DMEM. For reference purposes (100% values), 1 ml aliquots of the original bacterial cell suspensions used in the adhesion assay were centrifuged, the cells resuspended in 200 μl trypsin/EDTA plus 200 μl PBS and then frozen and stored at ‑20°C until quantification of the bacteria. B. infantis ATCC 15697 bacterial suspensions were carefully washed twice with sterile phosphate-buffered saline (PBS; pH 7.3) and approximately 1 x 108 cells of each strain were incubated at 37°C, 5% CO2 for 2h with a monolayer of fully differentiated Caco-2 cells. All incubations were performed in biologically independent triplicates. After 2 hours of incubation, cell monolayers were gently washed three times with PBS, to remove unbound bacteria.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Caco-2 cell monolayers were gently washed three times with PBS and RNA was extracted from Caco-2 cells using the Trizol method according to the manufacturer’s instructions (Invitrogen, Carlsbad, CA).
Sequencing libraries of mRNA were prepared using the Illumina TruSeq RNA Sample Prep Kit v2
Pooled libraries were sequenced on an Illumina HiSeq 2000. Sequencing was run for 100 cycles, with read length of 100 bp (single reads).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
INFANTIS_HMO2
|
| Data processing |
Reads were demultiplexed and imported into CLC-Bio Genomics Workbench Version 5.5.1
The following criteria were used to filter the unique sequence reads: minimum length fraction of 0.9; minimum similarity fraction of 0.8; maximum number of two mismatches.
Reads were mapped to the human reference genome, build 37.1
Data were normalized by calculating the reads per kilo base per million mapped reads (RPKM=total exon reads/mapped reads in millions x exon length in kb) for each gene
Gene annotations were based on the Ensembl human genome assembly GRCh37.p11
Genome_build: Homo sapiens, Build 37.1
Supplementary_files_format_and_content: tab-delimited text files contain abundance measurements as RPKM and counts
|
| |
|
| Submission date |
Dec 08, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Danielle G Lemay |
| E-mail(s) |
[email protected]
|
| Organization name |
University of California Davis
|
| Department |
Genome Center
|
| Street address |
451 Health Sciences Dr
|
| City |
Davis |
| State/province |
CA |
| ZIP/Postal code |
95616 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE63950 |
RNA sequencing of CACO-2 cells incubated with bifidobacteria grown on human milk oligosaccharides. |
|
| Relations |
| BioSample |
SAMN03255146 |
| SRA |
SRX799360 |
