E-MEXP-3913 - Transcription profiling by array of human LHK2 lung adenocarcinoma side population and main population cells
Released on 2 May 2014, last updated on 23 July 2014
We used the commercially available amino-allyl RNA amplification Kit ver,2 (High Yield Type) (SIGMA-ALDRICH). Purified total RNA (3 ﾵg) was reverse-transcribed to generate double-stranded cDNA using an oligo dT T7 promoter primer and reverse transcriptase. Next, cRNA was synthesized using T7 RNA polymerase, which simultaneously incorporated Cy3- or Cy5-labeled cytidine triphosphate. During this process, the samples of SP cells were labeled with Cy5, whereas the non-SP cells were labeled with Cy3 as control cells. Quality of the cRNA was again checked using the Nano Drop. Cy3-labeled cRNA and Cy5-labeled cRNA were combined and then fragmented in a hybridization cocktail (SIGMA-ALDRICH). Then the labeled cRNAs were hybridized to a 60-mer probe oligonucleotide microarray and incubated for 20 h ours at 50 ﾰC. The fluorescent intensities were determined by a Genepix 4000B Microarray Scanner (Axon, US).
transcription profiling by array, co-expression, dye swap design, self vs self design
Ectopically expressed variant form of sperm mitochondria-associated cysteine-rich protein augments tumorigenicity of the stem cell population of lung adenocarcinoma cells. Takahashi A, Hirohashi Y, Torigoe T, Tamura Y, Tsukahara T, Kanaseki T, Kochin V, Saijo H, Kubo T, Nakatsugawa M, Asanuma H, Hasegawa T, Kondo T, Sato N. PLoS One 8(11):e69095 (2013), PMID:24244262