Cloning and expression of a rat prothrombin cDNA

Abstract

A rat liver cDNA library was screened for a prothrombin gene using a cDNA coding human prothrombin as a probe. Twelve plaques identified by first screening were further screened. Two plaques were selected for sequencing. The DNA sequence of the gene coding rat prothrombin have been determined. It consistes of 1950 nucleotide and encodes an ORF of 1851 nucleotide, 22 bp noncoding regions at 5$^\prime$ end and 97 bp noncoding regions at 3$^\prime$ end that do not have poly A tail and poly A attachment signal, AATAAA. It shows 80\% homology with human prothrombin. Comparative analysis of region having considerable sequence homology between human and rat prothrombin suggests that the functional and structural domains and post translational modification sites in rat prothrombin can be mapped. The signal peptide, the cleavage sites, $\gamma$-carboxylated region, glycosylaton attachment sites and the cleavage sites by factor Xa in rat prothrombin can be presumed by comparative analysis. Rat prothrombin cDNA was expressed in E. coli as a fusion protein using pGEX-3X vector system.