| Ornithine decarboxylase: Expression and regulation in rat brain and in transgenic mice | ||
|---|---|---|
| Prev | Chapter 4. Materials and Methods | Next |
Expression levels of wild-type and D233V-ODC mRNAs in C55.7 cells were measured using Northern blot analysis. Total RNA from cultured cells was isolated by acid guanidinium thiocyanate-phenol-chloroform extraction (Chomczynski & Sacchi 1987). RNA gel electrophoresis and transfer onto nitrocellulose were performed by standard techniques (Sambrook et al. 1989). Blots were probed with a labelled 1691 bp SalI-PvuII fragment of the mouse ODC cDNA and autoradiographed. The autoradiography films were scanned using a laser densitometer (Computing Densitometer Model 300A, Molecular Dynamics, Sunnyvale, CA, USA). ODC mRNA levels in different samples were normalized by comparing them to the levels of 28S rRNA determined from the same blots.