As previously described (23 (link)), 5-6 weeks old, female DBA2/N mice (Envigo) were inoculated with 5 x 104 CFU of SN250 by lateral tail vein injection. After 48 hrs, mice were euthanized, kidneys harvested, and placed directly into ice-cold RNA Later solution (n = 6). The kidneys were then flash frozen in liquid nitrogen and ground into a fine powder with liquid nitrogen. The resulting tissue powder is mixed with the ice-cold Trizol. The samples were placed on a rocker at room temperature (RT) for 15 min and further the cell debris were removed by centrifuged the samples at 10K rpm at 4°C for 10 min. Cleared Trizol was collected into a new 1.5 ml Eppendorf tube and 200 μl of RNase free chloroform was added. Tubes were shaken vigorously for 15s and kept at RT for 5 min. Further the samples were centrifuged at 12K rpm for 15 min at 4°C. The cleared aqueous layer then transferred to new 1.5 ml tube and RNA was further extracted following the Qiagen RNeasy kit protocol.
Dba2 n mice
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DBA/2N mice are an inbred mouse strain commonly used in research. They are genetically identical and exhibit reproducible physiological and behavioral characteristics. The DBA/2N strain is widely utilized across various fields of study, including immunology, neuroscience, and drug development.
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2 protocols using dba2 n mice
1
Kidney RNA extraction from DBA2/N mice
2
Intradermal Injection of C. albicans
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The mutants and their respective parent strains were grown overnight in YPD at 30˚C. Harvested cells were washed thrice with sterile phosphate-buffered saline (PBS) and counted with a hemocytometer. A 1:1 mixture of NEON- tagged reference strain and iRFP-tagged mutant strain was mixed to get a final count of 1 × 108 CFU/ml in PBS. The 5- to 6-week-old female DBA2/N mice (Envigo) used in these experiments were maintained on chlorophyll-free chow to minimize endogenous fluorescence. Prior to injections, the mice were anesthetized with isoflurane using SomnoSuite low flow anesthesia machine (Kent Scientific) and the hair on the ears was removed by chemical depilation. Aliquots of 1 × 106 CFU/ml (10 μl) of C. albicans cells containing equal volume of reference and mutant strains (1:1) were injected into the dorsal ear dermis of anesthetized mice with a 29G1/2 needle. A characteristic papule was observed at the site of injection, indicating a successful intradermal injection.
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