Pan_neuro_control
Drosophila melanogaster
Protocols: Nuclei dissociation from frozen drosophila heads. 200ul of homogenisation buffer (250mM Sucrose (Fisher scientific), 10mM Tris pH 8.0 (Sigma Aldrich), 25mM KCL (Applichem), 5mM MgCl2 (Sigma-Aldrich), 0.10% Triton x 100 (Sigma-Aldrich), 0.1% DTT (NEB), 1x Protease inhibitor (Thermo scientific), 0.5% Rnasin ribonuclease inhibitor (Promega), Nuclease free water (Invitrogen)) was added to an Eppendorf with 60 flash frozen heads. Motorised pestle was used to grind samples for 2 mins on ice. A further 800ul homogenisation buffer was added and transferred to a dounce pestle. 60 strokes in the tight pestle. Sample was filtered through a 35 μm cell strainer into a 5ml FACs tube. Transferred into a 1.5ml Eppendorf and centrifuged for 10 mins at 1,000 g. Discarded supernatant and re-suspended nuclei in 500ul resuspension buffer (0.5% BSA (Sigma-Aldrich), 0.5% Rnasin ribonuclease inhibitor (Promega), 1x PBS (gibco)). Filtered sample through a 40 μm Flowmi cell strainer into a 5ml FACs tube. Sample was then FACs sorted for Hoescht+ nuclei into a collection tube. After FACs sorting, collection tube was centrifuged for 10 mins at 1,000 g. The supernatant was discarded and the pellet re-suspended with 45.2 μl resuspension buffer. Nuclei were then ready to be loaded to the 10x Chromium Controller.  Nuclei dissociation from frozen drosophila heads. 200ul of homogenisation buffer (250mM Sucrose (Fisher scientific), 10mM Tris pH 8.0 (Sigma Aldrich), 25mM KCL (Applichem), 5mM MgCl2 (Sigma-Aldrich), 0.10% Triton x 100 (Sigma-Aldrich), 0.1% DTT (NEB), 1x Protease inhibitor (Thermo scientific), 0.5% Rnasin ribonuclease inhibitor (Promega), Nuclease free water (Invitrogen)) was added to an Eppendorf with 60 flash frozen heads. Motorised pestle was used to grind samples for 2 mins on ice. A further 800ul homogenisation buffer was added and transferred to a dounce pestle. 60 strokes in the tight pestle. Sample was filtered through a 35 μm cell strainer into a 5ml FACs tube. Transferred into a 1.5ml Eppendorf and centrifuged for 10 mins at 1,000 g. Discarded supernatant and re-suspended nuclei in 500ul resuspension buffer (0.5% BSA (Sigma-Aldrich), 0.5% Rnasin ribonuclease inhibitor (Promega), 1x PBS (gibco)). Filtered sample through a 40 μm Flowmi cell strainer into a 5ml FACs tube. Sample was then FACs sorted for Hoescht+ nuclei into a collection tube. After FACs sorting, collection tube was centrifuged for 10 mins at 1,000 g. The supernatant was discarded and the pellet re-suspended with 45.2 μl resuspension buffer. Nuclei were then ready to be loaded to the 10x Chromium Controller.  Using Chromium Single Cell Expression 3' Reagent Kits Using Chromium Single Cell Expression 3' Reagent Kits
ENA first public
2025-12-31
INSDC center name
King's College London
INSDC status
public
Submitter Id
E-MTAB-14766:Pan_neuro_control
broker name
ArrayExpress
collection date
not collected
common name
fruit fly
genetic modification
Control
geographic location (country and/or sea)
not collected
isolate
not applicable
organism part
brain
sample name
E-MTAB-14766:Pan_neuro_control
scientific_name
Drosophila melanogaster