Cloning and functional analysis of cDNA encoding the hamster Bcl-2 protein

We have cloned cDNA encoding hamster Bcl-2 protein from total RNA of CHO-9 cells by RT-PCR using oligonucleotide primers sharing homology with the seq uence of mouse and rat bcl-2. The fragments spanning the total coding regio n were cloned into pCR4-TOPO and sequenced for verification. The hamster bc l-2 cDNA has a size of 711 nucleotides and encodes a polypeptide of 236 ami no acids. Hamster Bcl-2 shares 95.8 and 88.6% similarity with mouse and hum an Bcl-2, respectively. Northern blot analysis revealed a single 7.5 kb bcl -2 transcript in hamster (CHO-9), mouse (BK4), and rat (H5) cells and a 8.5 kb bcl-2 mRNA in human (HeLa MR) cells. The bcl-2 cDNA (771 bp) was reclon ed into pcDNA3 and the recombinant construct was transiently transfected in to MGMT-deficient HeLa MR cells. Expression of hamster Bcl-2 rendered the c ells more resistant to MNNG-induced cytotoxicity, which is consistent with the anti-apoptotic function of Bcl-2. (C) 2000 Academic Press.