Transcriptome Profiling of Cellular Aging

After subconfluent culture, 5×10⁶–1×10⁷ UE6E7T-3 cells at PDL 80 (Stage I), PDL110 (Stage II), PDL219 (Stage III) and PDL270 (Stage IV) were harvested. Each cellular total RNA from the four samples was extracted using QIAGEN RNeasy Mini Kit (Qiagen K. K., Tokyo, Japan). RNA samples were stored at −80°C until use. Poly-A RNA was used to select mRNA using the SOLiD RiboMinus Kit (Life Technologies, Gaithersburg, MD, USA). Following rRNA depletion, poly-A RNA was used to synthesize cDNA using the SOLiD Whole-Transcriptome Analysis Kit (Life Technologies) [25 (link)]. DNA sequencing (longest 50bp-reads) was carried out using SOLiD 3 PLUS System (Life Technologies) as described previously [26 (link)].