Production of recombinant human BRCA1 encoded proteins in E coli

Abstract

Among the risk factors that cause breast cancer, heredity emerges as the major determinant. BRCAl is the first gene which was found to be associated with inherited early-onset breast cancer. BRCAl is spread over a 100 kb region on human chromosome 17q21 and consists of 22 coding exons which are transcribed into a 7.8 kb long mRNA. This mRNA is abundant in breast and ovarian tissues and encodes a polypeptide of 1863 amino acids. The exact cellular function of BRCAl remains to be elucidated. As there are many gaps of knowledge and many conflicts about the cellular functions of BRCAl, new points of views and technical approaches should be generated. As such studies require the presence of purified protein products, we aimed to express and purify BRCAl encoded proteins. In this study we cloned the BRCAl gene in four overlapping fragments into the pCR-Script Amp, sk (+) cloning vector and subcloned the carboxyl terminal into the pQE expression vector. The 73 kD gene product was purified by affinity chromatography.