Mus musculus
Protocols: Tissues were harvested at prescribed circadian time on the third day of constant darkness. Mice were killed by cervical dislocation and their brains were rapidly dissected under dim red light. Brains were sectioned in ice-cold diethyl pyrocarbonate (DEPC)-treated phosphate buffered saline,pH 7.4, with an oscillating tissue slicer to obtain an 800μm thick coronal slice containing the SCN. The 800μm coronal slice containing the SCN was placed on a dry ice-chilled glass slide and the SCN was microdissected using a sharp scalpel blade. Each sample represented a pool of SCN tissues from 3 mice. RNA extraction was performed using the RNeasy Micro Kit according the manufacturer's instructions. RNA samples were submitted to the Génome Québec Innovation Centre at McGill University (Montréal, Canada) for RNA-seq library preparation. Total RNA was quantified using a Nanodrop Spectrophotometer ND-1000 (NanoDrop Technologies) and its integrity was assessed on a 2100 Bioanalyzer (Agilent Technologies). Libraries were generated from 250 ng of total RNA as follows: mRNA enrichment was performed using the NEBNext Poly(A) Magnetic Isolation Module (New England BioLabs). cDNA synthesis was achieved with the NEBNext RNA First Strand Synthesis and NEBNext Ultra Directional RNA Second Strand Synthesis Modules (New England BioLabs). The remaining steps of library preparation were done using the NEBNext Ultra II DNA Library Prep Kit for Illumina (New England BioLabs). Adapters and PCR primers were purchased from New England BioLabs. Libraries were quantified using the Quant-iT™ PicoGreen® dsDNA Assay Kit (Life Technologies) and the Kapa Illumina GA with Revised Primers-SYBR Fast Universal Kit (Kapa Biosystems). Average size fragment was determined using a LabChip GX (PerkinElmer) instrument.
ENA-FIRST-PUBLIC
2019-03-18T17:03:31Z
ENA-LAST-UPDATE
2018-12-05T10:49:53Z
External Id
SAMEA5147954
INSDC center name
Department of Biology, University of Toronto Mississauga; Department of Cell & Systems Biology, University of Toronto
INSDC first public
2019-03-18T17:03:31Z
INSDC last update
2018-12-05T10:49:53Z
INSDC status
public
Submitter Id
E-MTAB-7496:CT18_KO5
age
40
broker name
ArrayExpress
common name
house mouse
developmental stage
adult
genotype
Vgat-cre;Sox2fl/fl
growth condition
3 days constant darkness
organism part
suprachiasmatic nucleus
sample name
E-MTAB-7496:CT18_KO5
scientific_name
Mus musculus
sex
mixed
strain
mixed C57/BL6 x 129/SvEv x FVB