|
| Status |
Public on Apr 05, 2021 |
| Title |
EM1214: Kidney tissue from mice (1214) without infection and fed with a standard chow |
| Sample type |
SRA |
| |
|
| Source name |
Kidney
|
| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6
tissue: Kidney
treatment: Mice without leptospira infection and fed with a standard chow
|
| Treatment protocol |
C57BL/6 mice were inoculated by using intraperitoneal challenge with L. interrogans. At 28 days after infection, Leptospira-infected mice received a secondary nephrotoxic stimulation by adenine in diet for 14 days.
|
| Growth protocol |
C57BL/6 female mice aged 7–8 weeks were used for this study. All animal experiments were handled under Animal Biosafety Level 2 conditions and followed all appropriate guidelines for the use of infected animals.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA of kidney tissues was extracted using the TRIzol reagent (Invitrogen, Carlsbad, CA).
The first step in the workflow involves purifying the poly-A containing mRNA molecules using poly-T oligo-attached magnetic beads. Following purification, the mRNA is fragmented into small pieces using divalent cations under elevated temperature. The cleaved RNA fragments are copied into first strand cDNA using reverse transcriptase and random primers. This is followed by second strand cDNA synthesis using DNA Polymerase I and RNase H. These cDNA fragments then have the addition of a single 'A' base and subsequent ligation of the adapter. The products are then purified and enriched with PCR amplification. We then quantified the PCR yield by Qubit and pooled samples together to make a single strand DNA circle (ssDNA circle), which gave the final library.
DNA nanoballs (DNBs) were generated with the ssDNA circle by rolling circle replication (RCR) to enlarge the fluorescent signals at the sequencing process. The DNBs were loaded into the patterned nanoarrays and pair-end reads of 100 bp were read through on the BGISEQ-500 platform for the following data analysis study. For this step, the BGISEQ-500 platform combines the DNA nanoball-based nanoarrays and stepwise sequencing using Combinational Probe-Anchor Synthesis Sequencing Method.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
BGISEQ-500 |
| |
|
| Description |
Control mice
|
| Data processing |
SNP & INDEL calling; Sequencing Reads Filtering: SOAPnuke: version：v1.5.2
Sequenced reads were mapped to Mus_musculus mm10 reference genome using HISAT2 v2.0.4.
To reconstruct transcripts using StringTie v1.0.4, and with genome annotation information we identify novel transcripts by using Cuffcompare(a tool of Cufflinks v2.2.1) and predict the coding ability of those new transcripts using CPC v0.9-r2
After novel transcript detection, we merge novel coding transcripts with reference transcripts to get complete reference, then we map clean reads to it using Bowtie2 v2.2.5, then calculate gene expression level for each sample with RSEM v1.2.12
We use DEseq2 algorithms to detect the DEGs
|
| |
|
| Submission date |
May 15, 2020 |
| Last update date |
Apr 05, 2021 |
| Contact name |
Li-Fang Chou |
| E-mail(s) |
d928209@gmail.com
|
| Phone |
+886-3-3281200
|
| Organization name |
Chang Gung Memorial Hospital, Linkou
|
| Department |
Department of Medical Research and Development
|
| Lab |
Kidney Research Center
|
| Street address |
5 Fu-Shing St., Kueishan
|
| City |
Taoyuan |
| ZIP/Postal code |
333 |
| Country |
Taiwan |
| |
|
| Platform ID |
GPL23479 |
| Series (1) |
| GSE150641 |
RNAseq of kidneys from a mice model of chronic leptospirosis infection and induced a secondary nephrotoxic injury by adenine feeding |
|
| Relations |
| BioSample |
SAMN14931667 |
| SRA |
SRX8347256 |
