cell line: TOLEDO cell type: Diffuse Large B cell Lymphoma genotype: Wild-type
Treatment protocol
All the cells were non-treated
Growth protocol
All the cells were grown in supplemented media (RPMI or Iscove)
Extracted molecule
total RNA
Extraction protocol
RNA from cell lines was isolated using the RNeasy Plus MIni kit (QIAGEN). RNA integrity was determined using the RNA 6000 Nano LabChip Kit on an Agilent 2100 Bioanalyzer (Agilent Technologies). Ten micrograms of total RNA was used to synthesize double-stranded cDNA using Superscript III (Invitrogen), but using a modified oligo-d(T) primer (Supplemental Table 2). The second strand was synthesized for 2 hours at 16°C in a reaction containing 30 μl of 5× second-strand buffer (Invitrogen), 90 μl of RNAse-free water, 3 μl dNTP (10 mM), 1 μl of BSA (1 μg/μl), 1 μl of E. coli DNA ligase (10 U/μl) (Invitrogen), 4 μl E. coli DNA polymerase I (10 U/μl) (Invitrogen), and 1 μl of RNAse H (2 U/μl) (Invitrogen). Finally, 2 μl of T4 DNA Polymerase (5 U/μl) (Invitrogen) was added and the reaction incubated at 16°C for another 10 minutes. Reactions were cleaned with QIAGEN’s QIAquick PCR Purification Kit
Label
Cy3/Cy5
Label protocol
Labeling was performed by NimbleGen Systems Inc., Reykjavik, Iceland, following their standard operating protocol. See www.nimblegen.com.
Hybridization protocol
Hybridization was performed by NimbleGen Systems Inc., Reykjavik,Iceland, following their standard operating protocol. See www.nimblegen.com.
Scan protocol
Scanning was performed by NimbleGen Systems Inc., Madison, WI USA, following their standard operating protocol. See www.nimblegen.com.
Description
2 biological replicates
Data processing
The raw data (.pair file) was subjected to quantile normalization (Bolstad et al. Bioinformatics 19(2):185), and log2 transformation as implemented in the NimbleScan software package, version 2.3.16 (Roche NimbleGen, Inc.).
