noninfected_m5
Protocols: RNA was extracted (see nucleic acid extraction protocol) from freshly extracted small intestinal epithelium (see below) from mice infected with Nippostrongylus brasiliensis and not (see treatment protocol), a library was created (see nucleic acid library construction protocol) and sequenced paired end 150 bp (see nucleic acid sequencing protocol) Epithelium from small intestine was isolated using slightly modified version of a published protocol (Sato 2013). In summary, the small intestine of mouse was washed in ice-cold PBS and opened longitudinally. A microscopy cover slip was used to gently scrape of excess mucus and villus before the intestine was cut into 2-4 mm pieces. The fragments were resuspended in 30 mL ice-cold PBS and pipetted up and down with a 10 mL pipette. Supernatant was discarded and this step was repeated 5-10 times until the supernatant was clear. The fragments were incubated in 2 mM EDTA in PBS at 4 C for 30 min with gentle rocking. The supernatant was removed, 20 mL of ice-cold PBS was added and the fragments were pipetted up and down. This washing step was repeated until the crypt fraction appeared as seen in a microscope. The 1-3 consecutive fractions were collected into a FCS coated 50 mL tube. References: Sato, Toshiro and Hans Clevers (2013). “Primary Mouse Small Intestinal Epithelial Cell Cultures”. In:Epithelial Cell Culture Protocols: Second Edition. Ed. by Scott H. Randell and M. Leslie Fulcher. Methods in Molecular Biology. Totowa, NJ: Humana Press, pp. 319-328.isbn: 978-1-62703-125-7.doi:10.1007/978-1-62703-125-7_19.527url:https://doi.org/10.1007/978-1-62703-125-7_19. Mice were infected with Nippostrongylus brasiliensis RNA was extracted with a column based RNA extraction kit such as Qiagen spin miniprep. Library preparation was done with NEB Next(R) Ultra(TM) RNA Library Prep Kit
ENA-FIRST-PUBLIC
2021-05-01T00:23:35Z
ENA-LAST-UPDATE
2020-06-03T20:20:23Z
External Id
SAMEA6872460
INSDC center name
Dept. of Clinical and Molecular Medicine Norwegian University of Science and Technology (NTNU)
INSDC first public
2021-05-01T00:23:35Z
INSDC last update
2020-06-03T20:20:23Z
INSDC status
public
Submitter Id
E-MTAB-9183:noninfected_m5
broker name
ArrayExpress
common name
house mouse
developmental stage
adult
genotype
wild type genotype
individual
G
organism part
epithelium of small intestine
sample name
E-MTAB-9183:noninfected_m5
scientific_name
Mus musculus
strain
C57BL/6