|
| Status |
Public on Jan 20, 2023 |
| Title |
Nlrx1 deficient BMDCs 6hrs post challenge Replicate 2 |
| Sample type |
SRA |
| |
|
| Source name |
BMDCs challenged with viable Aspergillus fumigatus conidia
|
| Organism |
Mus musculus |
| Characteristics |
cell type: Mouse bone marrow derived dendritic cells
time: 6hrs post challenge
genotype: Nlrx1 deficient
|
| Treatment protocol |
Cells were challenged with 5 X 10^5 viable Aspergillus fumigatus conidia (AF29) and incubated for various times at 37C in the presense of 5% CO2.
|
| Growth protocol |
Bone marrow was extracted from the tibia and femur of wildtype and nlrx1 deficient mice. Cells were differentiated using GM-CSF. Cells were grown for 6 days with a medium exchange on day 3. On day 6 cells were harvested and plated at a density of 5 X10^5 cells per well (24-well format)
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were lysed in Buffer RLT containing BME. RNA was immediately extracted following the RNeasy Mini Kit Protocol. DNAase digestion was performed. Samples were eluted in 30ul twice.
Total RNA with RIN ≥ 8.0, was converted into a strand-specific library using Illumina’s TruSeq Stranded mRNA HT Sample Prep Kit (Illumina, RS-122-2103), for subsequent cluster generation and sequencing on Illumina's NextSeq. The library was enriched by 13 cycles of PCR, validated using Agilent TapeStation and quantitated by qPCR. Individually indexed cDNA libraries were pooled and sequenced on NextSeq
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
N8
|
| Data processing |
Illumina Casava1.8 software used for basecalling.
Sequences were trimmed for residual adaptors and poor quality bases (sliding window 4, phred score 25) and length (> 36) using Trimmomatic-0.35. Non-paired reads were also filtered out.
Filtered paired end reads were alligned to mouse reference genome (GRCm38.94) using HiSAT2
Transcript Assemble and quantification of RNA-Seq was done via Stringtie, with output specific for DESeq2
DESeq2 analysis was conducted to determine normalized gene counts and differential expression (Padj < 0.05, log2-fold change > 0.5). Differential expression was conducted in a pair-wise manner between genotypes or treatments
Genome_build: GRCm38.94
|
| |
|
| Submission date |
Jan 20, 2020 |
| Last update date |
Jan 20, 2023 |
| Contact name |
Shiv D. Kale |
| E-mail(s) |
sdkale@vt.edu
|
| Phone |
540-231-3784
|
| Organization name |
Virginia Tech
|
| Department |
Biocomplexity Institute
|
| Street address |
1015 Steger Hall, Life Science Circle
|
| City |
Blacksburg |
| State/province |
VA |
| ZIP/Postal code |
24061 |
| Country |
USA |
| |
|
| Platform ID |
GPL19057 |
| Series (1) |
| GSE143964 |
Time course RNA-Seq analysis of wildtype and Nlrx1 deficient bone marrow derived monocytoid dendritic cells indicate decision tree in response to A. fumigatus viable conidia |
|
| Relations |
| BioSample |
SAMN13893297 |
| SRA |
SRX7584856 |
