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Sample GSM154233 Query DataSets for GSM154233
Status Public on Jul 05, 2007
Title DNA-EP-4HRS_1
Sample type RNA
 
Source name tibialis cranialis muscle, exercised 4 hrs after plasmid DNA and in vivo electro gene transfer. Replicate one.
Organism Mus musculus
Characteristics Strain: C57Bl/6, gender: female, 8 weeks old.
Biomaterial provider Laboratory of the Department of Oncology, 5405 University of Copenhagen at Herlev Hospital, Herlev Ringvej 75, DK-2730 Herlev, Denmark
Treatment protocol Plasmid DNA and in vivo Electro gene transfer:
Five micrograms of the plasmid phGFP-S65T, encoding the green fluorescent protein (GFP), plasmid was dissolved in 20 μl PBS and injected intramuscularly along the fibres into the tibialis cranialis muscle of anaesthetised C57Bl/6 mice using an insulin syringe. Plate electrodes with 4-mm gap were fitted around the hind legs and an electric field was applied. The muscle was pulsed with a combination of a high voltage pulse (100 μsec, 1000V/cm) followed by a long low voltage pulse (400 ms, 100V/cm) with 1s lag between the pulses.
The tibialis cranialis muscle, exercised 4 hrs after plasmid DNA and in vivo electro gene transfer.
Growth protocol Six to eight weeks old female C57Black/C mice (average weight 22 g, Taconic, Denmark) were kept under pathogen-free conditions at 22 °C in a 14/10 hrs light/dark cycle with food and water ad libitum. All animal experiments were conducted in accordance with the recommendations of the European Convention for the Protection of Vertebrate Animals used for Experimentation and after permission from the Danish Animal Welfare Committee.
Extracted molecule polyA RNA
Extraction protocol Four hrs, 48 hrs and 3 weeks after treatment the mice were euthanized and the muscles were removed and placed in 1 ml solution D (guandinium thiocyanat, sodium citrate, sarcosyl and mercaptoethanol) on ice. Muscle extracts were prepared by homogenising the muscles with a rotor-stator homogeniser, and RNA were extracted using the Chomczynski and Sacchi method and digested with DNase. Due to low yield of total RNA, mRNA were amplified yielding antisense RNA (aRNA) using the MessageAMP II aRNA kit (Ambion, Europe).
Label Biotin
Label protocol 5 μg aRNA was used to synthesize double stranded cDNA using Superscript® Choice System (Invitrogen, Denmark) with an oligo(dT) primer containing a T7 RNA polymerase promoter (GenSet, Evry, France). The cDNA was used as template for an IVT reaction to generate biotinlabelled antisense cRNA (BioArrayTM High Yield RNA Transcript Labelling Kit; Enzo Diagnostics, Farmingdale, NY, USA).
 
Hybridization protocol The biotinlabelled antisense cRNA was fragmentated at 94°C for 35 min in fragmentation buffer (40 mM Tris, 30 mM MgOAc,10 mM KOAc), the labelled cRNA were hybridised for 16 hrs to Affymetrix MOE 430 2.0 arrays (Affymetrix, Santa Clara, CA, USA).The arrays were washed and stained with phycoerythrin conjugated streptavidin using the Affymetrix Fluidics Station® 400.
Scan protocol The arrays were scanned in the Affymetrix GeneArray® scanner to generate fluorescent images, as described in the Affymetrix GeneChip protocol.
Description Nothing in line.
Data processing The cel image files (Affymetrix) were imported, pre-processed and analysed in DNA-Chip Analyser 2006 (http://www.dchip.org). The array files were normalised using the non-linear invariant set normalisation
method, choosing array (DNA-EP-48HRS) as baseline. Normalisation curves were visually inspected and none of the arrays were critical. In order to interpret the probe signal, model-based gene expression indexes
(MBEI) gene expression modelling was calculated using the PM/MM perfect match/ miss match) method. During the analysis array outliers were detected. None of the arrays exceeded 1.4% of array outliers
and the probe presence call was between 36.6% and 50.3%.
 
Submission date Jan 08, 2007
Last update date Aug 28, 2018
Contact name John R. Zibert
E-mail(s) johnrobertz@geh.regionh.dk
Phone +45 3977 7538
Fax +45 3965 7137
Organization name Copenhagen University Hospital Gentofte
Department Dep. of Dermatology
Street address Niels Andersens Vej 65, KA-1502
City Hellerup
ZIP/Postal code 2900
Country Denmark
 
Platform ID GPL1261
Series (1)
GSE6686 Gene expression profiles in skeletal muscle after gene transfer by electroporation
Relations
Reanalyzed by GSE119085

Data table header descriptions
ID_REF
VALUE Normalised expression values

Data table
ID_REF VALUE
AFFX-BioB-5_at 336.896
AFFX-BioB-M_at 400.145
AFFX-BioB-3_at 376.895
AFFX-BioC-5_at 719.62
AFFX-BioC-3_at 1008.73
AFFX-BioDn-5_at 2292.09
AFFX-BioDn-3_at 4299.16
AFFX-CreX-5_at 7468.4
AFFX-CreX-3_at 6717.46
AFFX-DapX-5_at 33.8848
AFFX-DapX-M_at 115.824
AFFX-DapX-3_at 86.3608
AFFX-LysX-5_at 33.9987
AFFX-LysX-M_at 12.4586
AFFX-LysX-3_at 18.9518
AFFX-PheX-5_at 17.6444
AFFX-PheX-M_at 14.9849
AFFX-PheX-3_at 62.2845
AFFX-ThrX-5_at 38.8361
AFFX-ThrX-M_at 14.7928

Total number of rows: 45101

Table truncated, full table size 849 Kbytes.




Supplementary file Size Download File type/resource
GSM154233.CEL.gz 3.9 Mb (ftp)(http) CEL
Processed data included within Sample table

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