|
| Status |
Public on Apr 17, 2008 |
| Title |
Gene expression after infection with Y. enterocolitica pYV+, sublethal |
| Sample type |
RNA |
| |
|
| Source name |
CD11b+ cells of the spleen, infected with Y. enterocolitica pYV+, sublethal
|
| Organism |
Mus musculus |
| Characteristics |
Age: 6-8 weeks old
Gender: female
Strain: C57BL/6 mice
infected with Y. enterocolitica strain WA-314 (pYV+) for one day
|
| Treatment protocol |
Mice were infected with 5000 Y. enterocolitica WA-314 (pYV+), sublethal dose, for one day. After one day, five to ten mice per group were sacrificed by CO2 asphyxiation. The spleens were removed and placed in ice-cold HBSS (Ca2+ and Mg2+ free Hanks’ balanced salt solution [HBSS; BioWhittaker,Walkersville, MD.] supplemented with 2% FCS [HyClone, Logan, Utah] and 10 mM HEPES buffer). The spleens were forced with a 5 ml syringe pestle through a 100 µm-pore nylon mesh cell strainer (Falcon; BD Biosciences). Red blood cells were lysed from spleen samples by incubating the cell suspensions for 5 min at room temperature in erythrocyte lysis buffer (170 mM Tris, 160 mM NH4Cl, pH 7,4) followed by two washes in ice-cold HBSS. CD11b+ cells were purified by positive selection using MACS® CD11b MicroBeads (Miltenyi Biotec) according to the manufacturer´s instructions. To ensure enrichment values of 96-98% purity, the magnetic separation was performed twice. Cell viability and number were assessed by trypan blue exclusion.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from CD11b+ cells using the RNAeasy mini-kit following the manufacturer´s instructions (Qiagen, Hilden, Germany). Generation of fragmented cRNA was performed as recently described [38] and used for hybridization onto MG-U74Av2 from Affymetrix (Affymetrix, High Wycombe, UK). Genechips were washed, stained with streptavidin–phycoerythrin and read using a GeneChip Scanner 2500 (Affymetrix) at the IZKF microarray facility, Tübingen.
|
| Label |
Biotin
|
| Label protocol |
Standard procedure; Affymetrix protocol
|
| |
|
| Hybridization protocol |
Standard procedure; Affymetrix protocol
|
| Scan protocol |
Standard procedure; Affymetrix protocol
|
| Description |
CD11b+ cells of infected mice; Mice were infected with a sublethal dose of Yersinia enterocolitica WA-314 (pYV+) for one day
|
| Data processing |
Affymetrix ExpressionStat 5.0
|
| |
|
| Submission date |
Apr 16, 2008 |
| Last update date |
Apr 16, 2008 |
| Contact name |
Erwin Bohn |
| E-mail(s) |
[email protected]
|
| Phone |
00-49-70712981525
|
| Fax |
00-49-7071294972
|
| Organization name |
University of Tuebingen
|
| Department |
Institutefor Medical Microbiology and Hygiene
|
| Street address |
Elfriede-Aulhorn-Strasse 6
|
| City |
Tuebingen |
| ZIP/Postal code |
72076 |
| Country |
Germany |
| |
|
| Platform ID |
GPL81 |
| Series (1) |
| GSE11189 |
IFN-g counteracts YopH mediated immune evasion in Yersinia enterocolitica infection in mice |
|
