Northern, Western, enzymatic and correction studies. (A) Northern analysis of total RNA extracted from wild-type and Mut knockout livers. The mutase message is apparent in the wild-type sample and absent in the Mut mutant liver. Actin hybridization after stripping and reprobing shows the RNA to be intact and equally reactive. (B) Western blotting using anti-mutase antisera reveals a band of ~80 kd in the wild-type liver extracts that is completely absent from the mutant liver extracts. (C) [1-14C] propionic acid incorporation, with and without vitamin B12, in various cell lines, expressed as percent of wild-type activity. The wild type activity varied between days and ranged from 5.0–15.0 nmol C14 propionate/mg protein/18 hrs. Error bars surround the standard deviation from triplicate measurements. The corrected Mut null murine cell line (labeled MUTO + MCM LENTI) shows restored propionate flux when compared with the Mut null and GFP-transduced cell lines.
Chandler et al. BMC Medical Genetics 2007 8:64 doi:10.1186/1471-2350-8-64