regulated to 0.3-fold or less in mature oligodendrocytes were identified as genes required for the maintenance of OPC status. Fig. 3A and B illustrate diagram-matically the 4348 up-regulated and 1808 down- regulated genes in mature oligodendrocytes com-pared to OPCs as analyzed by three different algorithms. In addition, a total of 32 up-regulated and 19 down-regulated genes were selected from the intersection of the three algorithms (Fig. 3A, B). To confirm the accuracy of the microarray screen, we searched the expression profile of well-charac-terized genes required for the oligodendrocyte differentiation, such as mpz, plp1b, mag, cldnk and flj13639, and found that the expression levels of these genes were highly elevated in mature oligodendrocytes (Fig. 3C), indicating that our microarray analysis was successful. Validation of microarray data by quantitative RT-PCR (qRT-PCR) To validate the microarray data, we performed a qRT-PCR analy- sis of selected genes which ex-hibited highly elevated expression levels in the differentiated oligodendrocytes. We selected seven Fig. 4. Validation of microarray data by qRT-PCR. Selected target genes were evaluated by qRT-PCR using the RNA purified from the sorted OPCs and differentiated oligodendrocytes. (A) qRT-PCR results were normalized to β-actin, and data are presented as relative induction of differentiated oligodendrocytes compared with OPCs. Myelin basic protein (mbp) was used as a positive control. (B) Fold changes determined by qRT-PCR and microarray analysis are presented. http://dx.doi.org/10.5607/en.2011.20.2.85 Screening of the Myelination Factor genes not previously known to be involved in oligo-dendrocyte differentiation. The expression levels for bhlhe40 (basic helix- loop-helix family, member e40, NM_212679), dlg1 (discs, large (Drosophila) homolog 1, NM_199526), rbp2a (retinol binding protein 2a, cellular, NM_153004), rock2b (rho-associated, coiled-coil containing protein kinase 2b, NM_001093747), sepp1a (selenoprotein P, plasma, 1a, NM_178297), zgc:110190 (NM_001020570) and zgc:112160 (NM_001017724) were evaluated by qRT-PCR using RNA purified from the sorted OPCs and mature oligodendrocytes. Myelin basic protein (mbp) was used as a positive control, and the expression levels of the selected genes are presented as the relative induction in differentiated oligo- dendrocytes compared to OPCs. Consistent with the microarray profiles, qRT-PCR demonstrated that the expression of these seven genes was highly elevated in differentiated oligodendrocytes (Fig. 4), suggesting that all seven genes may be involved in oligoden- drocyte differentiation. Basic helix-loop-helix family member e40 (bHLHB2, also known as Dec1/Eip1/Sharp2/Stra13/Clast5) is a novel cAMP- inducible bHLH protein family member. Stra13 is required for the proliferation and dif-ferentiation of nerve cells. Stra13 controls growth arrest, and represses the expression of cell proli-feration associated genes through an HDAC1- independent mechanism (Sun and Taneja, 2000). Dec1 is expressed in the nuclei of endothelial, glial and neuronal cells in brain tissue (Preusser et al., 2005). Since the bHLHB2 expression level is elevated in differentiated oligodendrocytes, we hypothesize that bHLHB2 plays a role in oligo-dendrocyte differentiation by regulating the OPC cell cycle. Discs-large homolog 1 (Dlg1) is a multi- domain scaffolding pro- tein that is involved in protein trafficking in polarized cells and membrane addition in Drosophila (Lee et al., 2003; Gorczyca et al., 2007). Dlg1 is expressed in myelin-forming Schwann cells, and enriched in Schwann cell paranodes where myelin outfoldings preferentially arise. In-terestingly, dlg1 interacts with kinesin 13B (kif13B) and Sec8, which play a central role in oligoden-drocyte membrane formation by regulating the re-cruitment of vesicles that transport myelin proteins, suggesting that dlg1 function may be required for oligodendrocyte differentiation (Bolis et al., 2009). Retinol binding protein 2a (Rbp2a/CRBP) is a cyto- plasmic retinol-specific binding protein and is expressed in the lateral ganglionic eminences (LGE), where a second wave of OPCs originate in the telencephalon (Kessaris et al., 2008). Retinol is naturally converted into retinoic acid which regulates the timing of oligodendrocyte differentiation in vivo (Barres et al., 1994). Therefore, we hypothesize that Rbp2a may play a role in oligodendrocyte development together with retinol. Rho-GTPase family proteins play important roles in coordinating www.enjournal.org 89