|
| Status |
Public on Apr 25, 2024 |
| Title |
LymphNode_MutantToxin_1 |
| Sample type |
RNA |
| |
|
| Source name |
Inguinal and axillary lymph nodes
|
| Organism |
Mus musculus |
| Characteristics |
tissue: Inguinal and axillary lymph nodes
treatment: C. difficile glucotransferase null mutant toxin B2
Sex: Female
age: 8 weeks
strain: C57Bl/6NCr
|
| Treatment protocol |
TcdB2 and D270N were expressed in Bacillus megaterium (MoBiTec, Göttingen, Germany) and purified by nickel affinity chromatography (GE Life Sciences, Boston, MA) as previously described. Purity and integrity were confirmed by SDS-PAGE and each batch of TcdB2 was tested for toxicity using a CHO cell killing assay. Mice were injected s.c. with 10 ng TcdB2 in PBS (controls were given PBS or 10 ng D270N in PBS).
|
| Growth protocol |
Female C57BL/6 mice (C57BL/6NCr) at 8 weeks of age were purchased from Charles River Laboratories (Bethesda, MD, Cat # model 027). All animal procedures were approved by the Institutional Animal Care and Use Committee at the University of Oklahoma Health Sciences Center (OUHSC) (original protocol 20-020-AHI, renewed as 22-082-CHI). Procedures were performed under inhalational anesthesia using a 4% isoflurane / 96% medical air mixture dispersed through a precision vaporizer. This study was performed in accordance with the recommendations of the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The aLNs and iLNs were isolated after 7 days. Lymph nodes were placed immediately in DNA / RNA Shield (Zymo Research, Irvine, CA). RNA was purified from the lymph nodes using Zymo Quick-RNA Miniprep Plus Kit (Zymo Research, Irvine, CA). Purified RNA was stored at -80°C until needed.
|
| Label |
N/A
|
| Label protocol |
N/A
|
| |
|
| Hybridization protocol |
N/A
|
| Scan protocol |
RNA analysis was performed by the OUHSC core facility using a NanoString nCounter SPRINT profilerTM (NanoString, Seattle, WA) in conjunction with the nCounter Myeloid Innate Immunity PanelTM.
|
| Description |
D versus P baseline: no significant genes from 654 hits. T versus D: 3 significant genes from 654 hits. Myeloid Innate Immunity Panel with total of 754 genes.
D1
|
| Data processing |
Data were analyzed with nSolver 4.0 Advanced Analysis SoftwareTM (Version 2.0.134, NanoString, Seattle, WA). Analysis was performed by normalizing the raw transcript counts utilizing negative synthetic sequences to account for background noise and positive synthetic sequences to account for technical variations.
Data summarizes the log2 fold change, raw p values, and adjusted p values (Benjamin-Yekutieli method) for each two-way comparison in the experiment: TcdB2 versus PBS, TcdB2 versus D270N, and D270N versus PBS.
|
| |
|
| Submission date |
Apr 11, 2024 |
| Last update date |
Apr 25, 2024 |
| Contact name |
Kaylee M Norman |
| E-mail(s) |
Kaylee-norman@ouhsc.edu
|
| Phone |
19189987829
|
| Organization name |
University of Oklahoma Health Sciences Center
|
| Department |
Microbiology and Immunology
|
| Lab |
Lang Lab
|
| Street address |
940 Statond L Young Blvd
|
| City |
Oklahoma City |
| State/province |
OK |
| ZIP/Postal code |
73104 |
| Country |
USA |
| |
|
| Platform ID |
GPL25266 |
| Series (1) |
| GSE263792 |
Clostridioides difficile Toxin B subverts germinal center formation and antibody recall responses by stimulating a drug-treatable CXCR4-dependent mechanism |
|
