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Sample GSM678325 Query DataSets for GSM678325
Status Public on Jul 18, 2011
Title ZF-TF 83, 2nd infection
Sample type RNA
 
Source name MCF7
Organism Homo sapiens
Characteristics retroviral transduction: zinc finger transcription factor 83
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 ZF-TF.83.b3
fulvestrant sensitive MCF7 clone retroviraly induced with zinc finger transcription factor 83, 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
E-mail(s) wittner.ben@mgh.harvard.edu
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
ID_REF
VALUE MAS5 signal intensity, batch-corrected

Data table
ID_REF VALUE
1007_s_at 2265.994377
1053_at 229.8457526
117_at 52.96950628
121_at 347.6478321
1255_g_at 10.0824939
1294_at 125.113245
1316_at 42.58028847
1320_at 80.20418172
1405_i_at 14.11944002
1431_at 17.59446716
1438_at 153.8302312
1487_at 231.5674791
1494_f_at 71.55978697
1552256_a_at 246.0490263
1552257_a_at 437.3906666
1552258_at 39.09662013
1552261_at -2.033742354
1552263_at 31.73280445
1552264_a_at 187.1614744
1552266_at 8.923728068

Total number of rows: 54675

Table truncated, full table size 1214 Kbytes.




Supplementary file Size Download File type/resource
GSM678325.CEL.gz 5.1 Mb (ftp)(http) CEL
Raw data provided as supplementary file
Processed data included within Sample table

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