Nanodrop nd 2000 spectrophotometer

Prior to osteogenic induction, SMSCs were collected and used as control group at day 0 for comparison between different breeds and tissues. Subsequently, at 1, 7, 14, and 21 days of cultivation, SMSCs in ODM and in parallel in CCM were collected. Total RNA of cells at each time point were isolated using TRI reagent (Sigma-Aldrich, Taufkirchen, Germany) and the RNeasy kit (Qiagen, Hilden, Germany) according to the manufacturer’s protocols. Both the quantity and purity of total RNA were measured using a NanoDrop ND-2000 spectrophotometer (Peqlab, Erlangen, Germany), while the integrity of RNA was determined via 1% agarose gel electrophoresis. All RNA samples were stored at − 80 ℃. Total RNA (500 ng) per sample was used for cDNA synthesis by the Affymetrix GeneChip WT PLUS Reagent Kit (Affymetrix, Santa Clara, CA, USA). The cDNA was fragmented and biotin-labeled via the Affymetrix GeneChip WT Terminal Labeling Kit (Affymetrix, Santa Clara, CA, USA), and then was hybridized on a genome-wide snowball array (Affymetrix, Santa Clara, CA, USA), containing 47,880 probe sets describing 17,964 annotated genes base on the Sscrofa11 genome [SNOWBALLs520824F]. After staining and washing, Affymetrix GCOS 1.1.1 software was used for scanning and processing the array data.