Figure 4
Pair wise analysis by a modified t-test of SAM and the validation of the cDNA microarray data. (A) The scatter plot of the observed relative difference (di) versus the expected relative difference dE(j). The solid line indicates the line for d(i) = dE(j), where the observed relative difference is identical to the expected relative difference. The dashed lines are drawn at a distance Δ = 0.758 from the solid line. The 474 potentially significant genes for Δ = 0.758 are indicated by red circles (255 up-regulated genes) and green circles (219 down-regulated genes) from the 14k dataset (14,077 genes). (B) The raw expression values (log2) of the 474 differentially expressed genes for the SCK cells, as compared to the control for the Choi-CK cells, are shown rank-ordered according to the fold change score. The triplicate expression ratios (the Cy5/Cy3 ratio) between the experimental samples (SCKCy5-1, -2, and -3) and the reference (Choi-CKCy3-1, -2, and -3) were repeated after swapping dye (SCKCy3-1, -2, and -3 versus Choi-CKCy5-1, -2, and -1). (C) Northern blot analysis of the LGALS1, TGFBI, CES1, LDHB, UCHL1, ASPH, VDAC1, VIL2, CCND2, S100P, CALB1, MAL2, GPX1 and ANXA8 gene expressions for the validation of the cDNA microarray data. The total RNA extracts from the JCK, Cho-CK, Choi-CK and SCK cells were fractionated by electrophoresis on 1% agarose gels that contained formaldehyde, and the proteins were then transferred to membranes. The blots were hybridized overnight with 2 × 106 cpm/ml of each cDNA probe labeled with [P32]dCTP (NEN) by randompriming; they were then washed and next exposed to X-Omat AR film (Kodak) at -70℃. The blot was stripped and subsequently rehybridized with a probe for 18S cDNA as a loading control (lower).
