2025-06-30
ENA first public
ERC000011
ENA-CHECKLIST
University of Heidelberg Center for Molecular Biology (ZMBH)
INSDC center name
public
INSDC status
ERS23984762
SRA accession
E-MTAB-15025:ovary_SN334_GD76
Submitter Id
GD76
age
ArrayExpress
broker name
mixture of gonadal cell type
cell type
not collected
collection date
white-tufted-ear marmoset
common name
Protocols: Frozen tissue was homogenized on ice in a buffer containing sucrose, KCl, MgCl2, Tris-HCl (pH 8), IGEPAL, DTT, Murine RNase Inhibitor, SUPERase-In, and cOmplete Protease Inhibitor Cocktail. The tissue was disrupted by trituration and/or using a micropestle. After a brief incubation, unlysed tissue debris was removed by low-speed centrifugation (100g for 1 minute at 4°C). The supernatant was then centrifuged at 400g for 4 minutes to separate the nuclei (pellet) from the supernatant. Nuclei were washed once or twice in the homogenization buffer and resuspended in a storage buffer containing sucrose, KCl, MgCl2, Tris-HCl (pH 8), Murine RNase Inhibitor, SUPERase-In, and cOmplete Protease Inhibitor Cocktail. If needed, the nuclei were filtered using 40 µm Flowmi strainers. Nuclei concentration was determined by staining with Hoechst DNA dye or propidium iodide and counting on a Countess II FL Automated Cell Counter. Following the nuclei count, approximately 15,000 nuclei from each nuclei suspension were used for generating snRNA libraries. For early marmoset gonad samples (GD74 and GD80), around 5,000 nuclei were use for library generation due to the small size of the gonads at these stages. RNA was extracted from the cytoplasm extracts or nuclei suspensions by mixing them with RLT buffer (supplemented with 40 mM DTT) and 100% ethanol in a 2:7:5 ratio. The RNA was then purified using the RNeasy Micro Kit from Qiagen. The quality of the extracted RNA was assessed using a Fragment Analyzer (Advanced Analytical), and all samples had RNA quality numbers (RQN) above 8. Single-cell barcoding and library preparation were performed using Chromium Single Cell 3' Reagent Kits (v3 chemistry) and the Chromium Controller instrument (10x Genomics), following the manufacturer's protocols. The cDNA was amplified using 12 PCR cycles. The Qubit Fluorometer (Thermo Fisher Scientific) was used to quantify the libraries, and the average fragment size was assessed using a Fragment Analyzer.
description
embryo stage
developmental stage
wild type genotype
genotype
not collected
geographic location (country and/or sea)
MAR Embryo 2  GD76 070918 G
individual
not applicable
isolate
Callithrix jacchus
organism
Callithrix jacchus
organism
ovary
organism part
Callithrix jacchus
scientific_name
female
sex
ovary_SN334_GD76
title