2025-12-31
ENA first public
ERC000011
ENA-CHECKLIST
King’s College London
INSDC center name
public
INSDC status
ERS22949382
SRA accession
E-MTAB-14766:Pan_neuro_control
Submitter Id
ArrayExpress
broker name
not collected
collection date
fruit fly
common name
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
description
Control
genetic modification
not collected
geographic location (country and/or sea)
not applicable
isolate
Drosophila melanogaster
organism
Drosophila melanogaster
organism
brain
organism part
Drosophila melanogaster
scientific_name
Pan_neuro_control
title