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Sample GSM738128 Query DataSets for GSM738128
Status Public on May 22, 2012
Title Zic3 NPC 3
Sample type RNA
 
Source name ZiNPC
Organism Homo sapiens
Characteristics cell type: ZiNPC derived from BJ1 fibroblasts
gender: male
cell line: BJ1
treatment: Zic3, KLF4, OCT4 and SOX2
Biomaterial provider SCIL
Treatment protocol The coding region of the mouse Zic3 gene was amplified by RT-PCR and cloned in pMIG-IRES-GFP plasmid (Addgene). The pMXs plasmids encoding for OCT4, SOX2 and KLF4 were purchased from Addgene. pMXs and pMIG-based retroviral vectors were transfected individually along with the viral packaging genes gag-pol (Addgene) and VSVG (Addgene) into 293T cells (ATCC) using Fugene HD reagent (Roche) according to the manufacturer's directions. Human BJ1 cells (Lonza) were seeded at 1.5 x 105 cells/well in 6-well plates (Sigma) without feeders. Cells were transduced with Zic3, KLF4, OCT4 and SOX2 retroviral vector containing supernatants mixed equally. To achieve maximum transduction efficiency, cells were transduced twice with the viral supernatant. Four days after transduction, the transduced cells were reseeded at 1.5 x 105 cells per 100-mm dish containing MEF and cultured with hESC medium. ZiNPC were picked in 15-20 days post transduction.
Growth protocol mTESR medium
Extracted molecule total RNA
Extraction protocol Rneasy (Qiagen)
Label biotin
Label protocol RNA concentration and purity were determined spectrophotometrically using the Nanodrop ND-1000 (Nanodrop Technologies) and RNA integrity was assessed using a Bioanalyser 2100 (Agilent). Using the Ambion WT Expression Kit, per sample, an amount of 100 ng of total RNA spiked with bacterial poly-A RNA positive controls (Affymetrix) was converted to double stranded cDNA in a reverse transcription reaction. Next the sample was converted and amplified to antisense cRNA in an in vitro transcription reaction which was subsequently converted to single stranded sense cDNA. Finally, samples were fragmented and labeled with biotin in a terminal labeling reaction according to the Affymetrix WT Terminal Labeling Kit.
 
Hybridization protocol A mixture of fragmented biotinylated cDNA and hybridisation controls (Affymetrix) was hybridised on Affymetrix  GeneChip Human Gene 1.1 ST Array Plate followed by staining and washing in the GeneTitan® Instrument (Affymetrix) according to the manufacturer's procedures.
Scan protocol To assess the raw probe signal intensities, chips were scanned using the GeneTitan® HT Array Plate Scanner (Affymetrix).
Description BJ1 fibroblast transduced with OCT4, SOX2, KLF4 and Zic3
Data processing RMA-normalized expression values as obtained with the xps package (version 1.10.2) of BioConductor
 
Submission date Jun 07, 2011
Last update date May 23, 2012
Contact name Rekin's Janky
E-mail(s) Nucleomics.Bioinformatics@vib.be
Organization name VIB
Department Nucleomics Core
Street address Herestraat 49 Box 816
City Leuven
ZIP/Postal code B-3000
Country Belgium
 
Platform ID GPL11532
Series (1)
GSE29770 Transduction of human fibroblasts with Zic3 combined with OCT4, SOX2 and KLF4 induces stable neural progenitor cell lines

Data table header descriptions
ID_REF
VALUE RMA-normalized expression values as obtained with the xps package (version 1.10.2) of BioConductor

Data table
ID_REF VALUE
7896736 6.684664475
7896738 3.792178605
7896740 3.242181754
7896742 10.19692173
7896744 6.223561561
7896746 9.114861513
7896748 8.997128661
7896750 4.864027625
7896752 10.74965996
7896754 8.399478745
7896756 4.577773221
7896759 7.168652063
7896761 5.998904037
7896779 7.539228617
7896798 6.076841245
7896817 5.80634726
7896822 7.92372562
7896859 4.266081884
7896861 3.573180603
7896863 4.681083766

Total number of rows: 28204

Table truncated, full table size 547 Kbytes.




Supplementary file Size Download File type/resource
GSM738128_hyb10340.CEL.gz 4.8 Mb (ftp)(http) CEL
Processed data included within Sample table

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