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Sample GSM678321 Query DataSets for GSM678321
Status Public on Jul 18, 2011
Title Control, 2nd infection
Sample type RNA
Source name MCF7
Organism Homo sapiens
Characteristics retroviral transduction: control
Treatment protocol 293T cells were seeded at 5 x 105 cells per 10 cm culture dish, transfected with 3.2 μg of ZT-TF p65 activator library DNA and 2.4μg and 0.8μg of PMD-MLV and PMD-G plasmid DNA using Fugene 6 according to the manufacture’s protocol and retroviral supernatant harvested 48 hrs later. MCF7-R73 cells (6 x105 per 10 cm plate) were infected with 1.33 x 105 ZF-TF retrovirions in media containing polybrene. Two days post-infection the cells were exposed to puromycin (0.4mg/ml), and three days later were subjected to continuous combined puromycin and fulvestrant (100nM) selection for an additional 42 days. Retroviral DNA was recovered by PCR, subcloned into the pDONOR plasmid using Gateway technology (Invitrogen), and subject to DNA sequencing that revealed 46 unique ZF-TF arrays. The 46 ZF-TFs were re-cloned into the retroviral destination plasmid using Gateway technology, 46 ZF-TF retroviral supernatants generated, and MCF-R73 cells infected and subjected to puromycin and fulvestrant selection. Six unique ZF-TFs conferred drug resistance in the presence of continuous fulvestrant exposure. A control MCF7 cell line (MCF7-R238) was generated by infecting MCF7-R73 cells with a retrovirus bearing the p65 activation domain only. T47D cells were infected with the six unique ZF-TF bearing retroviruses and subjected to fulvestrant exposure as described for the MCF7 cells.
Growth protocol MCF7-R73 cells, a clonal isolate derived from the MCF7 cell line were kindly provided by Dr. Toshi Shioda (MGH, Charlestown, MA) and maintained in a xenoestrogen controlled environment as described in Coser KR, Wittner BS, Rosenthal NF, Collins SC, Melas A, et al. (2009) Antiestrogen-resistant subclones of MCF-7 human breast cancer cells are derived from a common monoclonal drug-resistant progenitor, Proc Natl Acad Sci U S A 106: 14536-14541
Extracted molecule total RNA
Extraction protocol QIAGEN RNeasy protocol and reagents for extraction of total RNA
Label biotin
Label protocol Affymetrix One-Cycle Target Labeling and Control Reagents (Part#900493) + Enzo BioArray High Yield RNA Transcript Labelling Kit (Part#42655)
Hybridization protocol Microarray = Affymetrix GeneChip Human Genome U133 Plus 2.0. Affymetrix Hybridization Oven= 645. Affymetrix Fluidics Station= 450. Fluidics Protocol= EukGE-WS2
Scan protocol Affymetrix GeneChip Scanner= 3000-7G. Controller= GCOS
Description Control.b3
fulvestrant sensitive MCF7 clone retroviraly induced with control plasmid encoding only the NF-kappaB p65 activation domain, 2nd infection
Data processing MAS5 algorithm as implemented by call.exprs function of Bioconductor simpleaffy package version 2.14.05, 2% trimmed mean set to 100. The expression values were corrected for batch effect on a per-probe-set basis as follows. For each batch N other than the first, we added to the expression values of batch N the difference between the average expression value on the first batch and the average expression value on batch N.
Submission date Feb 22, 2011
Last update date Jul 18, 2011
Contact name Ben S. Wittner
Organization name Massachusetts General Hospital
Department Center for Cancer Research
Lab Lawrence
Street address 149 13th Street
City Boston
State/province MA
ZIP/Postal code 02129
Country USA
Platform ID GPL570
Series (1)
GSE27444 Expression data from zinc-finger-transcription-factor-induced Fulvestrant-Resistant MCF7 Cell Lines

Data table header descriptions
VALUE MAS5 signal intensity, batch-corrected

Data table
1007_s_at 1302.122482
1053_at 341.4139271
117_at 25.02024642
121_at 278.0552124
1255_g_at 10.92379877
1294_at 103.2557388
1316_at 27.19158342
1320_at 9.562268542
1405_i_at 14.5658346
1431_at 6.267159917
1438_at 126.9967003
1487_at 223.8043416
1494_f_at 67.94251217
1552256_a_at 324.7159006
1552257_a_at 397.6305488
1552258_at 42.36984356
1552261_at 11.92251374
1552263_at 27.08774874
1552264_a_at 194.5696368
1552266_at 17.16708091

Total number of rows: 54675

Table truncated, full table size 1214 Kbytes.

Supplementary file Size Download File type/resource
GSM678321.CEL.gz 5.0 Mb (ftp)(http) CEL
Raw data provided as supplementary file
Processed data included within Sample table

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