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| Status |
Public on Sep 21, 2017 |
| Title |
Control- 6hr-1 |
| Sample type |
SRA |
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| Source name |
NHBE cells, negative control, 6 hr
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| Organism |
Homo sapiens |
| Characteristics |
tissue: lung
cell type: NHBE
infection: Control
time: 6hr
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| Treatment protocol |
Cells were sub-cultured into six-well tissue culture dishes at a concentration of 1 x 106 cells per well and allowed to adhere overnight. The cells were washed twice with DPBS and cultured in a final volume of 1.5mL BEGM media without addition of the provided gentamicin/amphotericin-B component to avoid antifungal effects. Nine individual samples were prepared for each treatment by addition of 0.5 x 106 A. fumigatus spores or left untreated (control). Cells were incubated at 37°C and 5% CO2. Samples were collected at 2, 6, and 12 hrs., following initial stimulation. Culture supernatant was collected and debris was removed by centrifugation before storing at -80°C. The cells were washed twice with DPBS before RNA extraction.
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| Growth protocol |
Normal human bronchial epithelial (NHBE) cells were grown in bronchial epithelial growth media (BEGM) with SingleQuot (Lonza) supplements and growth factors (bovine pituitary Extract [BPE], hydrocortisone, human epidermal growth factor [hEGF], epinephrine, transferrin, insulin, retinoic acid, triiodothyronine, and bovine serum albumin – fatty acid free [BSA-FAF]) as recommended by the manufacturer. Cells were cultured in 75 cm2 tissue culture flasks at 37°C and 5% CO2 until reaching 80% confluence.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was collected by addition of 1mL TRIzol (Invitrogen Carlsbad, CA) directly to the cell culture plate and gentle agitation using a cell lifter. RNA extraction was carried out per manufacturer’s instructions followed by clean-up using the Qiagen RNeasy Kit (Qiagen, Valencia, CA) with on-column DNase digestion to remove residual genomic DNA. RNA was eluted in 50μl of RNase-free water and integrity (A260/A280) and concentration were assessed using a NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, DE). For each time point, equal amounts of RNA from three treatment replicates were pooled to one sample for a total of three samples per time point in duplicates. A control replicate pool was also generated from each of the three time points (2, 6, and 12 hrs.) in the same manner. RNA samples were then stored at -80°C until further analysis.
library was constructuted at Iowa State University prior to data processing as described at http://www.dna.iastate.edu/nextgensequencing.html
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
Sample 3
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| Data processing |
reads aligned using Bowtie 2 2.0.0 and Tophat 2.0.8 to hg19
transcripts were assembled using Cufflinks 2.0.1
we compared assmbled transcripts using Cufflinks 2.0.1
used Cuffmerge (found in Cufflinks 2.0.1) to merge transcripts
Cuffdiff (found in Cufflinks 2.0.1) was used to find differentially expressed genes
Genome_build: hg19
Supplementary_files_format_and_content: 2hr.txt: CuffDiff output of comparison of 2 hr samples control with A. fumigatus treated.
Supplementary_files_format_and_content: 6hr.txt: CuffDiff output of comparison of 6 hr samples control with A. fumigatus treated.
Supplementary_files_format_and_content: 12hr.txt: CuffDiff output of comparison of 12 hr samples control with A. fumigatus treated.
Supplementary_files_format_and_content: 2hr_sig.txt: Statistical signifcant values (described as yes in the signficance columnn of 2 hr file).
Supplementary_files_format_and_content: 6hr_sig.txt: Statistical signifcant values (described as yes in the signficance columnn of 6 hr file).
Supplementary_files_format_and_content: 12hr_sig.txt: Statistical signifcant values (described as yes in the signficance columnn of 12 hr file).
Supplementary_files_format_and_content: 2hr_sig_final.txt: selection of only those genes found to be statistically significant in (2 hr sig) but removal of the genes meeting the condition q-value=p-value=0.
Supplementary_files_format_and_content: 6hr_sig_final.txt: selection of only those genes found to be statistically significant in (6 hr sig) but removal of the genes meeting the condition q-value=p-value=0.
Supplementary_files_format_and_content: 12hr_sig_final.txt: selection of only those genes found to be statistically significant in (12 hr sig) but removal of the genes meeting the condition q-value=p-value=0.
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| Submission date |
Oct 01, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Reinhard Laubenbacher |
| E-mail(s) |
laubenbacher@uchc.edu
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| Organization name |
University of Connecticut Health Center
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| Department |
Center for Quantitative Medicine MC 6029
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| Street address |
263 Farmington Avenue
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| City |
Farmington |
| State/province |
CT |
| ZIP/Postal code |
06030 |
| Country |
USA |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE61955 |
Host transcriptome analysis of Aspergillus fumigatus infection in Airway Epithelial Cells |
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| Relations |
| BioSample |
SAMN03085685 |
| SRA |
SRX718203 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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