2025-07-25
ENA first public
ERC000011
ENA-CHECKLIST
Department of Internal Medicine, Yonsei University, College of Medicine
INSDC center name
public
INSDC status
ERS25081659
SRA accession
E-MTAB-15253:TSC22D1_500_4
Submitter Id
ArrayExpress
broker name
K1 PTC cell line
cell line
thyrocyte
cell type
not collected
collection date
human
common name
Protocols: The K1 papillary thyroid cancer (PTC) cell line was seeded in a 10-cm dish and harvested 24 hours post-transfection. Cells were maintained at less than 80% confluency throughout the experiment. Specific protocol description is not applicable to the present study. Cells were cultured in RPMI medium supplemented with 10% fetal bovine serum (FBS) and Normocin® (InvivoGen, Cat# ant-nr-1). Transfection was performed 24 hours after seeding. Mycoplasma contamination was routinely tested using the Myco-Read™ Mycoplasma Detection Kit (PCR) (Biomax, Cat# SMD0172). Transfection was performed using X-tremeGENE™ HP DNA Transfection Reagent (Roche, Cat# XTGHP-RO) according to the manufacturer’s protocol. The TSC22D1 (NM_006022) Human Tagged ORF Clone (Origene, Cat# RC223210) was transfected at two concentrations: 200 ng (low) and 500 ng (high). The pCMV6-Entry Mammalian Expression Vector (Origene, Cat# PS100001) was used as an empty vector control. Specific protocol description is not applicable to the present study. Total RNA was extracted using TRIzol reagent. For tissue samples, frozen tissue was finely minced, ground under liquid nitrogen, and immediately homogenized in 1 mL TRIzol. Cell samples were lysed by pipetting in TRIzol, followed by 5 minutes of incubation at room temperature. Phase separation was performed by adding 250 µL chloroform (or 100 µL 1-bromo-3-chloropropane), vortexing, incubating for 10 minutes, and centrifuging at 12,000 rpm for 10 minutes at 4°C. The aqueous phase was transferred, mixed with 500 µL isopropanol, incubated, and centrifuged at 14,000 rpm for 20 minutes at 4°C to pellet RNA. The pellet was washed twice with 75% ethanol, air-dried, and resuspended in 15–30 µL DEPC-treated water. RNA purity was confirmed by spectrophotometry, with acceptable A260/A280 ratios near 1.8. RNA libraries were prepared using the Illumina TruSeq Stranded mRNA Sample Prep Kit. mRNA was isolated with poly-T magnetic beads, fragmented, and converted to cDNA, followed by end repair, adapter ligation, and PCR enrichment.
description
N/A
developmental stage
thyroid cancer
disease
BRAFV600E
genotype
not collected
geographic location (country and/or sea)
not applicable
isolate
Homo sapiens
organism
Homo sapiens
organism
thyroid gland
organism part
Homo sapiens
scientific_name
TSC22D1 overexpression (high level)
stimulus
TSC22D1_500_4
title