Fig. 6.

Real-time PCR of myelinating cocultures. SCs infected with non-targeting shRNA (shCTRL) or shRNA against MLCK (shMLCK), were used to establish myelinating cocultures. RNA was isolated from a total of 12 cultures at the indicated time points and the relative increase of myelin protein mRNAs (P0, MBP and MAG), as well as Krox-20 and laminin mRNAs was measured by quantitative RT-PCR. Relative mRNA levels for each protein, normalized to actin (or L19), were calculated as indicated in Material and Methods. A significant increase in the relative amount of mRNA is observed for all proteins after 14 days (d) in coculture, reaching a plateau by 21 days. In shMLCK cultures (filled circles) the relative increase of mRNA is significantly higher (***P<0.001; **P<0.01, two-way ANOVA, Bonferroni post-test) than that of control cultures (open squares). The only exception was for the relative levels of MBP mRNA at 14 days, which was higher in control cultures.