2024-06-01
ENA first public
ERC000011
ENA-CHECKLIST
SAMEA114411254
External Id
Sun Yat-sun University Zhongshan Ophathlamic Center
INSDC center name
2023-09-22T10:27:26Z
INSDC last update
public
INSDC status
ERS16399918
SRA accession
E-MTAB-13364:Sample 1
Submitter Id
4 to 15
year
age
ArrayExpress
broker name
not collected
collection date
crab-eating macaque
common name
Protocols: Total 8 monkeys (4-15 years old, males only, Macaca fascicularis) were used in the current project. All procedures complied with the NIH Guide for the Care and Use of Laboratory Animals approved by Sun Yat-sen University. Following 2 weeks of adaptive housing in the institutional animal facility, animals were anesthetized with a lethal dose of sodium pentobarbital. Animals were transcardially perfused with 2 liters of artificial cerebrospinal fluid (ACSF, in mM: 125 NaCl, 25 NaHCO3, 1 NaH2PO4, 2 KCl, 25 D-Glucose, 2 CaCl2, 1 MgCl2, 5 sodium l-ascorbate, 3 sodium pyruvate, 0.01 taurine, 2 thiourea, 1 kynurenic acid, 0.1 DL-AP5, and 1x10-3 tetrodotoxin). The flow of perfusion fluid was adjusted so that the perfusion finished within approximately 40 minutes. The brain was removed from the skull immediately and a block of tissue containing the LIP was cut off. The position of LIP referred to the atlas of the rhesus monkey brain (Saleem and Logothetis, 2012). The tissue block was kept in the ACSF and snap frozen. The brain was removed from the skull immediately and a block of tissue containing the LIP was cut off. The single-nuclei suspensions were prepared following previously described methods (Krishnaswami et al. 2016), with some modifications. Briefly, the frozen brain tissues were lysed gently in a glass dounce homogenizer with 2 ml ice-cold Lysis solution, consisting of 250 mM sucrose, 25 mM KCl, 5 mM MgCl2, 10mM Tris buffer pH 8.0, 1mM DTT, 40 U/µL100x protease inhibitor, RNAse Inhibitor, and 0.1% Triton X-100. The lysate was kept on ice for 5 minutes and then filtered through a 40 μm filter (BD Falcon, 352350) to eliminate cell clumps. The nuclei pellets were collected by centrifuging the lysate at 500 g for 5 minutes and discarding the supernatant. The nuclei were then purified with density gradient centrifugation buffer solutions (25% and 29% iodixanol solution in 250 mM sucrose, 25 mM KCl, 5mM MgCl2, and 10 mM Tris buffer, pH 8.0) and centrifuged for 30 minutes at 13500g. The nuclei pellets were washed and stored at a resuspension buffer consisting of 1% UltraPure BSA (Sigma, B2064), 0.2 U/µL Rnase Inhibitor, in Rnase-free PBS (Gibco, C1001050BT). Nuclei suspensions with a viability of > 90% were used for snRNA-seq library and sequencing. Following the protocols provided by 10X Genomics Chromium platform.
description
adult
developmental stage
wild type genotype
genotype
not collected
geographic location (country and/or sea)
pool of 8 individuals
individual
not applicable
isolate
Macaca fascicularis
organism
Macaca fascicularis
organism
brain
organism part
lateral intraparietal cortex
sampling site
Macaca fascicularis
scientific_name
male
sex
Sample 1
title