3 10.4 – 9.1 9.1   2.3 2.4   212107_s_at DEAH ARS-1620 (Asp-Glu-Ala-His) box polypeptide 9 DHX9 – - – - – - – - – 212917_x_at RecQ protein-like (DNA helicase Q1-like) RECQL 10.6 10.7 – 9.5 9.6   2.2 2.3 – 212918_at RecQ protein-like (DNA helicase Q1-like) RECQL – - – - – - – - – 213520_at RecQ protein-like

4 RECQL4 – - – - – - – - – 213647_at DNA2 DNA replication helicase 2-like (yeast) DNA2L 8.6 8.7 8.7 10.2 10.2 10.2 -3.0 -2.8 -2.8 213878_at similar to EX 527 CG10721-PA LOC642732 – - – - – - – - – 221686_s_at RecQ protein-like 5 RECQL5 – - – - – - – - – Three different methods for data normalization using ACTB, GAPDH, and Affymetrix U-133A housekeeping selleck inhibitor genes, respectively were utilized. Table 3 Expression of DNA polymerase alpha. Probe set Description Gene symbol PT3 Non-PT3 Fold Differences       ACTB GAPDH U133-A ACTB GAPDH U133-A ACTB GAPDH U133-A 204441_s_at Polymerase (DNA directed), alpha 1 POLA1 – - – - – - – - – 204835_at Polymerase (DNA

directed), alpha 1 POLA1 11.7 11.8 11.8 10.1 10.1 10.1 2.9 3.1 3.1 Three different methods for data normalization using ACTB, GAPDH, and Affymetrix U-133A housekeeping genes, respectively were utilized. Comparison SPTLC1 of Normalization Techniques Based on the number of transcripts identified as differentially expressed, the three techniques used to normalize the array data could be ordered by the number of

genes identified as differentially expressed as follows: GAPDH (1869 probe sets) > ACTB (1781 probe sets) > U-133A (1478 probe sets). Although the three array normalization methodologies differed in the number of genes defined as down- or up-regulated in expression in PT3 compared to PT1 and NK cell lines, all identified the same 7 up-regulated genes (PCNA, POLD1, RFC3, RFC4, RFC5, RPA1, and RPA2) except RPA1 in normalization using HG-U133A housekeeping genes (Table1). This finding suggested that these seven genes were clearly differentially over-expressed in PT3 versus PT1 and NK cell lines. Verification of microarray results by real-time quantitative PCR As we did the microarray analysis using a single mRNA isolation/cDNA probe analysis, we needed to verify the transcriptional over-expression of these seven genes by real-time quantitative PCR. To determine the optimum amount of cDNA template in initial experiments, we performed undiluted, 1:10 diluted, and 1:100 diluted cDNA template in parallel.