biostudies-arrayexpressJoanna JurczakHomo sapienshttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-16674The aim of this study was to identify stably expressed genes suitable for RT-qPCR normalization in an in vitro model of human airway epithelium (AE). RNA sequencing was performed to characterize global gene expression profiles across AE cultures representing various sexes, AE origin (bronchial or nasal) and stages of epithelial differentiation. Strand-specific poly(A)-selected libraries were prepared using the TruSeq Stranded mRNA protocol and sequenced at Macrogen Europe BV (Amsterdam, Netherlands) on the Illumina NovaSeq 6000 platform in paired-end mode (2 × 150 bp), generating approximately 100 million reads per sample. The deposited files include raw paired-end FASTQ data generated from 2 primary and 27 human AE cells differentiated in vitro in ALI (air-liquid interface) culture. These data were used for transcriptome-wide assessment of gene expression stability and selection of candidate reference genes for downstream validation.biostudies-arrayexpressSample Collection - Primary human airway epithelial (AE) cells were obtained from healthy, non-smoking Caucasian donors. Nasal epithelial cells (NECs) were collected by nasal brushing from volunteers without acute airway inflammation within the preceding 6 weeks (except one commercially acquired sample). Bronchial epithelial cells (BECs) were obtained commercially (Epithelix or PromoCell). Primary cells were expanded and differentiated in vitro under air–liquid interface (ALI) conditions. Samples were collected at defined stages of differentiation (pre-ALI, ALI day 0, 3, 6, 9, 12, 21). Two samples (one male, one female NEC donor) represent RNA from uncultured primary AE cells isolated directly after collection. In total, 29 samples from multiple donors (NEC and BEC) were subjected to RNA sequencing.Nucleic Acid Extraction - Total RNA was isolated from cultured or primary airway epithelial cells using TRI Reagent (Thermo Fisher Scientific) according to the manufacturer’s instructions, followed by DNase treatment using TURBO DNA-free™ Kit (Thermo Fisher Scientific).Library Construction - Strand-specific poly(A)-selected libraries were prepared using the Illumina TruSeq Stranded mRNA Library Prep Kit according to the manufacturer’s protocol.Sequencing - Libraries were sequenced on the Illumina NovaSeq 6000 platform in paired-end mode (2 × 150 bp), generating approximately 100 million reads per sample.OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesSequence Alignment - Sequencing reads were aligned to the GRCh38 reference genome using STAR ver. 2.7.10a, with GENCODE transcript database ver. 48 (comprehensive, primary regions).Data Transformation - Read counts for individual genes were obtained using featureCounts from the Subread package ver. 1.6.3 and normalized using the TPM method.UnknownTranscriptomicsGenomicsProteomicsIllumina NovaSeq 6000RNA-seq of coding RNAHomo sapiensJoanna JurczakEwa ZiętkiewiczAdam UstaszewskiHanna Przystałowska-MaciołaRoman JaksikZuzanna Bukowy-BieryłłofalseNew validated reference genes for gene expression studies in the differentiating human airway epitheliumThe aim of this study was to identify stably expressed genes suitable for RT-qPCR normalization in an in vitro model of human airway epithelium (AE). RNA sequencing was performed to characterize global gene expression profiles across AE cultures representing various sexes, AE origin (bronchial or nasal) and stages of epithelial differentiation. Strand-specific poly(A)-selected libraries were prepared using the TruSeq Stranded mRNA protocol and sequenced at Macrogen Europe BV (Amsterdam, Netherlands) on the Illumina NovaSeq 6000 platform in paired-end mode (2 × 150 bp), generating approximately 100 million reads per sample. The deposited files include raw paired-end FASTQ data generated from 2 primary and 27 human AE cells differentiated in vitro in ALI (air-liquid interface) culture. These data were used for transcriptome-wide assessment of gene expression stability and selection of candidate reference genes for downstream validation.2026-05-11T00:00:00Z2026-05-13T14:11:12.632Z2026-02-24T23:11:22.231ZE-MTAB-16674ERP189571EFO_0002944EFO_0004170EFO_0004917EFO_0005518EFO_0003816EFO_0003738EFO_0004184