Quantifying Osteogenic Marker Expression

The expression of select osteogenic marker genes was assessed by quantitative PCR (qPCR), essentially as previously described, with some minor modifications [35 ,67 (link)]. The Taqman primers (from Allied Biosystems, ThermoFisher) were: ALPL (Mm00475834_m1), RUNx2 (Mm00501584_m1) and Osteonectin/SPARC/BM40 (Mm00486332_m1). Quantitative PCR (qPCR) was performed in a RealPlex Real-Time PCR System (Eppendorf, Enfield, CT) with fast thermal cycling, as previously described [35 ]. The level of expression of each gene was normalized to the level of expression of a common standard housekeeping gene, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and a relative expression set against the control at 1G in MM. Fold change was calculated via the comparative CT method (2^−ΔΔCT) [68 (link)].