Cloning and characterization of two alleles of the murine extracellular superoxide dismutase gene

Abstract

We have recently documented the existence of a second allele of ecSOD in mice. Thus far, this allele was only found in the 129P3/J strain. It is characterized by two point mutations leading to amino acid changes as well as a 10 bp deletion from the 3′ UTR. We have also shown that the phenotype is profoundly affected by the genotype. In order to obtain a tool to investigate the differences in the properties as well as the posttranscriptional regulation of expression of the two alleles we now describe the creation and characterization of stably transfected CHO-K1 cell lines expressing either of these alleles. CHO-K1 cells were chosen because they do not express endogenous ecSOD and are easy to transfect. We demonstrate that the transfected cells secrete substantial amounts of glycosylated ecSOD, detected by Western blot analyses, ConA–Sepharose affinity chromatography and activity measurements.