2025-08-01
ENA first public
ERC000011
ENA-CHECKLIST
Department of Internal Medicine, Division of Gastroenterology and Hepatology, University of Michigan Medical School
INSDC center name
public
INSDC status
ERS25081764
SRA accession
E-MTAB-15263:Colon 88 long-term 2D transwell epithelium day 7 (early)
Submitter Id
33
year
age
ArrayExpress
broker name
88-CA
cell line
Heterogeneous
cell type
not collected
collection date
human
common name
Protocols: Cells were gently scraped off the transwell membrane with a mini cell scraper in 200 µL of cold DPBS and transferred to a cryovial containing 1.3 mL of cold DPBS with a cut 0.1% BSA coated 200 µL pipette tip and centrifuged at 500 x g for 3 minutes at 4°C. Excess DPBS was gently removed from the cell pellet and the pellet was immediately flash frozen in liquid nitrogen. All samples were stored in liquid nitrogen until nuclei isolation. Single nuclei were isolated from flash frozen cell pellets using Chromium Nuclei Isolation kit (10x Genomics 1000493) and protocol for Single Cell Gene Expression and Chromium Fixed RNA Profiling (10x Genomics CG000505 Rev A) with added RNase inhibitor (2 U/mL, Roche 3335402001) throughout the isolation.  Because this protocol is designed for isolation of nuclei from whole tissue, we modified the initial lysis steps for gentler dissociation of the cell pellets by triturating the frozen cell pellet in cold lysis buffer 15 times with a 1 mL pipette and omitting the use of a pellet pestle. This was followed by a 5 minute incubation in the lysis buffer on wet ice with additional trituration at 2.5 minutes and 5 minutes of the incubation.  Isolated nuclei were counted and loaded into the 10x Chromium to create single-nuclei droplets targeting 5,000 nuclei. Single-cell libraries were prepared using the Next GEM Single Cell 3' Gel Bead kit v3.1 (10x genomics PN-1000147) according to the manufacturer's instructions by the University of Michigan Advanced Genomics Core.
description
adult
developmental stage
normal
disease
wild type genotype
genotype
not collected
geographic location (country and/or sea)
not applicable
isolate
Homo sapiens
organism
Homo sapiens
organism
ascending colon
organism part
Homo sapiens
scientific_name
female
sex
Colon 88 long-term 2D transwell epithelium day 7 (early)
title