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Sample GSM533875 Query DataSets for GSM533875
Status Public on Apr 20, 2010
Title si DOT1L + 9hr Wnt
Sample type RNA
 
Channel 1
Source name HEK293T human embryonic kidney cells
Organism Homo sapiens
Characteristics cell line: HEK293T
transfection: si-cDOT1L
induction: +Wnt3A, 9 hours
Treatment protocol Control or Wnt3A conditioned medium was added to the HEK293T cells and incubate for 9 hours before RNA extraction
Growth protocol Ls174T or HEK293T cells were seeded on 6-well plates and transfected with control/human b-catenin/MLLT10/DOT1L/BRG1/P300 15nM siRNA for 72 hours
Extracted molecule total RNA
Extraction protocol Total RNA extracted using RNeasy mini kit (Qiagen) following manufacturer's instructions
Label Cy3
Label protocol 2µg of total RNA were primed with 1.2 µl of T7 promoter primer at 65°C for 10 min together with control spike A/B, then reversed transcribed at 40°C for 2hrs in the presence of MMLV RTase (Agilent), and dNTP mix. It was then followed by the cRNA transcription at 40°C for 2hrs in the presence of T7 RNA polymerase, inorganic pyrophosphatase, NTP mix, DTT, PEG and Cy3/Cy5-labeled-CTP.
 
Channel 2
Source name Human reference RNA
Organism Homo sapiens
Characteristics reference: Human reference RNA
Treatment protocol Control or Wnt3A conditioned medium was added to the HEK293T cells and incubate for 9 hours before RNA extraction
Growth protocol Ls174T or HEK293T cells were seeded on 6-well plates and transfected with control/human b-catenin/MLLT10/DOT1L/BRG1/P300 15nM siRNA for 72 hours
Extracted molecule total RNA
Extraction protocol Total RNA extracted using RNeasy mini kit (Qiagen) following manufacturer's instructions
Label Cy5
Label protocol 2µg of total RNA were primed with 1.2 µl of T7 promoter primer at 65°C for 10 min together with control spike A/B, then reversed transcribed at 40°C for 2hrs in the presence of MMLV RTase (Agilent), and dNTP mix. It was then followed by the cRNA transcription at 40°C for 2hrs in the presence of T7 RNA polymerase, inorganic pyrophosphatase, NTP mix, DTT, PEG and Cy3/Cy5-labeled-CTP.
 
 
Hybridization protocol The synthezied cRNA was then purified with RNeasy mini kit (Qiagen). 825ng of the corresponding Cy3 and Cy5 cRNA was fragmentated at 60°C for 30 min in the presence of blocking agent and fragmentation buffer (Agilent), then combined with 55ul 2X hybridization buffer (Agilent) and applied to 4X44k human expression microarrays enclosed in Agilent SureHyb-enabled hybridization chambers. After hybridization, slides were washed sequentially
Scan protocol Scanned on an Agilent G2505 DNA microarray scanner.
Description 293T cells transfected with si-DOT1L, induced with Wnt3A medium for 9 hours.
Data processing Agilent Feature Extraction Software (v 8.5.1.1) was used for background subtraction and LOWESS normalization.
 
Submission date May 07, 2010
Last update date May 07, 2010
Contact name Vivian Li
E-mail(s) vivian.li@crick.ac.uk
Organization name The Francis Crick Institute
Lab Stem Cell and Cancer Biology
Street address 1 Midland Road
City London
ZIP/Postal code NW1 1AT
Country United Kingdom
 
Platform ID GPL4133
Series (1)
GSE21367 The Leukemia-associated Mllt10/Af10- Dot1l, dedicated b-catenin coactivators essential for intestinal homeostasis

Data table header descriptions
ID_REF
VALUE normalized log10 ratio Cy5/Cy3

Data table
ID_REF VALUE
1 -3.030489837e-001
2 0.000000000e+000
3 0.000000000e+000
4 0.000000000e+000
5 0.000000000e+000
6 0.000000000e+000
7 0.000000000e+000
8 0.000000000e+000
9 0.000000000e+000
10 0.000000000e+000
11 0.000000000e+000
12 6.842527361e-001
13 -9.688667856e-001
14 -3.716938664e-002
15 2.937586728e-001
16 2.087899927e-001
17 4.013727934e-001
18 -3.856888323e-001
19 1.136687004e-001
20 -6.806592678e-002

Total number of rows: 45015

Table truncated, full table size 1022 Kbytes.




Supplementary file Size Download File type/resource
GSM533875.txt.gz 4.4 Mb (ftp)(http) TXT
Processed data included within Sample table

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