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Sample GSM2735953

Query DataSets for GSM2735953

Status

Public on Oct 22, 2017

Title

mock infected HAE-1

Sample type

SRA

Source name

mock HAE cells

Organism

Homo sapiens

Characteristics

tissue: human airway epithelium (HAE)-ALI

Treatment protocol

The differentiated HAE cells were infected by HBoV1 at a MOI of 1000 viral genome copies (vgc)/cell or mock infected

Growth protocol

Primary human airway epithelial cells were cultured on collagen-coated, semipermeable membrane inserts of 0.33 cm2, and then were polarized/differentiated at an ALI for 3-4 weeks. Briefly, in the first 2 to 3 days after 2 ×104 airway epithelial cells were seeded onto the Transwell insert, SAGM-H media were fed in both the apical and basolateral chambers of the insert to start the culture, then the SAGM-H media were aspirated from both chambers, and the cells were fed with 500 µl of PneumaCultTM-ALI medium (StemCell, Vancouver, BC, Canada) in the basolateral chamber. The medium was changed every 3-4 days. The ALI-cultured HAE took 3-4 weeks for full differentiation. We chose the cultures with a transepithelial electrical resistance (TEER) of over 1,000 Ω∙cm2, as determined with an epithelial Ohm-voltmeter (Millicell-ERS; EMD-Millipore, Billerica, MA), for subsequent HBoV1 infection.

Extracted molecule

total RNA

Extraction protocol

At 7 days post-infection, mock and HBoV1 infected-cell were collected and RNA samples were extracted using the miRNeasy Mini Kit by following the manufacturer’s instruction
The TruSeq Stranded Total RNA with Ribo-Zero HMR library Prep Kit (#RS-122-2201, Illumina, San Diego, CA) was used to prepare the sequencing library from 1 μg of total RNA.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 2500

Data processing

Illumina bcl2fastq software was used for basecalling.
Mapping of RNA-seq reads, transcript assembly, and abundance estimation were conducted using Tuxedo Suite pipeline (TopHat v2.0.9/Cufflinks v2.2.1) and reported with Fragments Per Kilobase of exon per Million fragments mapped (FPKM)
Cuffdiff was used to calculate statistical significance changes to gene expression between HBoV1 and mock-infected HAE cells.
Genome_build: GRCh38.83
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample

Submission date

Aug 09, 2017

Last update date

May 15, 2019

Contact name

Jianming Qiu

E-mail(s)

jqiu@kumc.edu

Phone

9135884413

Organization name

University of Kansas Medical Center