title: 6M_CEREBELLUM
attributes.tag: BIOLOGICAL_REPLICATES
attributes.value: 2
attributes.tag: DISEASE_ONTOLOGY_CURIE
attributes.value: ncim:C0549184
attributes.tag: DISEASE_ONTOLOGY_URI
attributes.value: https://ncim.nci.nih.gov/ncimbrowser/ConceptReport.jsp?dictionary=NCI Metathesaurus&code=C0549184
attributes.tag: DONOR_AGE_UNIT
attributes.value: month
attributes.tag: DONOR_HEALTH_STATUS_ONTOLOGY_CURIE
attributes.value: ncim:C0549184
attributes.tag: DONOR_HEALTH_STATUS_ONTOLOGY_URI
attributes.value: https://ncim.nci.nih.gov/ncimbrowser/ConceptReport.jsp?dictionary=NCI Metathesaurus&code=C0549184
attributes.tag: DONOR_LIFE_STAGE
attributes.value: adult
attributes.tag: ENA first public
attributes.value: 2022-01-24
attributes.tag: ENA last update
attributes.value: 2022-01-24
attributes.tag: External Id
attributes.value: SAMEA11469594
attributes.tag: INSDC center alias
attributes.value: Center for Epigenomics Research, HKUST
attributes.tag: INSDC center name
attributes.value: center for epigenomics research, hkust
attributes.tag: INSDC first public
attributes.value: 2022-01-24T00:26:28Z
attributes.tag: INSDC last update
attributes.value: 2022-01-24T00:26:28Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: SAMPLE_ONTOLOGY_CURIE
attributes.value: uberon:0002037
attributes.tag: SAMPLE_ONTOLOGY_URI
attributes.value: http://purl.obolibrary.org/obo/UBERON_0002037
attributes.tag: Submitter Id
attributes.value: 6M_CEREBELLUM
attributes.tag: biological replicate
attributes.value: 2
attributes.tag: biomaterial provider
attributes.value: Leung lab, Hong Kong University of Science and Technology
attributes.tag: biomaterial type
attributes.value: Primary Tissue
attributes.tag: brain_region_name
attributes.value: Cerebellum
attributes.tag: brain_region_ontology
attributes.value: MA:0000198
attributes.tag: collection method
attributes.value: The animal was deeply anesthetized with Avertin (0.03 ml/g); the depth of anesthesia was assessed by the absence of deep tendon reflexes following a strong foot pinch. Once adequate anesthesia was achieved, the chest cavity was opened. The right atrium was clipped to allow exsanguination and the mouse was perfused through the left ventricle with PBS. Whole brain was obtained and cut into the left and right hemispheres. Each half was then dissected separately by eye. Hindbrain and cerebellum was first separated from the hemispheres, then cerebellum was obtained. The olfactory bulb was removed, along with brain stem regions until hippocampus was clearly visible. Hippocampus was obtained, and the remaining tissue was labelled as cerebral cortex. All collected tissue was snap frozen and stored at -80°C until nuclei extraction. For nuclei extraction, frozen tissue was added to 1 mL cold 1X homogenization buffer (320 mM sucrose, 0.1 mM EDTA, 0.1% NP40, 5 mM CaCl2, 3 mM Mg(Ac)2, 10 mM Tris pH 7.8, 16.7 μM PMSF, 167 μM β-mercaptoethanol, 1U/mL RNAse inhibitor in nuclease-free water) (modified from Corces et al., 2017). Tissue was dounced 10 strokes using loose pestle and 5 strokes using tight pestle (WHEATON® Dounce Tissue Grinder, 1 mL). Homogenate was filtered through a 40 μm cell strainer and spun down at 4°C, 1000g for 10 mins. Pellet was resuspended in 3mL of low sucrose buffer (0.32M sucrose, 10mM HEPES pH 8, 5mM CaCl2, 3mM Mg(Ac)2, 0.1mM EDTA, 1mM DTT, 0.2U/μL RNAse inhibitor in nuclease-free water) (modified from Matson et al., 2018). 12.5 mL of sucrose density buffer (1M sucrose, 10mM HEPES pH 8, 3mM Mg(Ac)2, 1mM DTT in nuclease-free water) was added slowly underneath the low sucrose buffer homogenate. The sucrose gradient was then centrifuged in a swing-bucket rotor at 4°C, 3200g for 20 min. The supernatant was decanted and pelleted purified nuclei was resuspended in buffer for sorting (0.2U/μL RNAse inhibitor, 1% BSA in PBS). Nuclei was stained with Hoechst at 2μg/uL.
attributes.tag: common name
attributes.value: house mouse
attributes.tag: donor age
attributes.value: 17
attributes.tag: donor id
attributes.value: 6M
attributes.tag: donor sex
attributes.value: male
attributes.tag: mouse id
attributes.value: 6M
attributes.tag: mouse_sacrificial_date
attributes.value: 2020-12-01
attributes.tag: nuclei_extraction_date
attributes.value: 2021-04-23
attributes.tag: sample name
attributes.value: 6M_CEREBELLUM
attributes.tag: strain
attributes.value: C57BL/6J
attributes.tag: tissue depot
attributes.value: Cerebellum
attributes.tag: tissue type
attributes.value: Cerebellum